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FOUNDED  BY  JOHN  D.  ROCKEFELLER 


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A  STUDY  OF  REDUCTION  IN  OENOTHERA 

RUBRINERVIS 


A  DISSERTATION 

SUBMITTED  TO  THE  FACULTY  OF  THE  OGDEN  GRADUATE  SCHOOL  OF 
SCIENCE  IN  CANDIDACY  FOR  THE  DEGREE  OF 
DOCTOR  OF  PHILOSOPHY 

(department  of  botany) 


BY 

REGINALD  RUGGLES  GATES 


Reprinted  from 

Botanical  Gazette,  Vol.  XLVI,  No.  i 
Chicago,  1908 


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Composed  and  Printed  By 
The  University  of  Chicago  Press 
Chicago,  Illinois,  U.  S.  A. 


VOLUME  XLVI 


NUMBER  I 


J- 

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Botanical  Gazette 


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JULY  jooA 


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A  STUDY  OF  REDUCTION '.IN  .OEiVO?T?BI?T 

RUBRINERVIS- 


CONTRIBUTIONS  FROM  THE  HULL  BOTANICAL  LABORATORY  III 

Reginald  Ruggles  Gates 

(with  plates  i-iii) 

The  present  contribution  is  a  statement  of  some  of  the  results 
obtained  in  the  cytological  study  of  Oenothera  Lamarckiana  and  its 
mutants.  Since  these  results  have  a  more  or  less  direct  bearing  on  a 
wide  range  of  fact  and  theory  in  various  fields,  their  full  discussion  is 
reserved  for  a  future  time.  The  facts  presented  in  this  communica¬ 
tion  will  be  taken  almost  wholly  from  the  study  of  O.  rubrinervis,  one 
of  the  most  vigorous  of  the  mutants.  Other  papers  will  be  presented 
later,  giving  the  further  evidence  upon  which  the  conclusions  of  this 
paper  rest,  and  attempting  to  indicate  their  bearing  on  the  general 
problems  of  cytology  and  variation  involved. 

Material 


The  plants  from  which  the  material  for  these  studies  was  obtained 
were  grown  from  pedigreed  seeds  of  DeVries,  the  purity  of  these 
cultures  being  further  proven,  in  some  cases,  by  carrying  on  the  pedi¬ 
gree  for  another  generation  before  collections  were  made.  The 
results  of  these  cultures,  which  are  still  being  carried  forward  to  later 
generations,  will  be  published  at  another  time  in  connection  with  an 
account  of  other  studies  on  variation  and  hybridization  in  Oenothera. 
In  this  way  it  is  hoped,  if  possible,  to  correlate  the  cytological  data 
with  the  work  in  hybridization  and  variation.  It  seems  to  the  writer 
that  only  by  thus  combining  cytological  with  experimental  studies  is  an 
explanation  of  the  peculiar  and  remarkable  phenomena  of  variation 
exhibited  by  the  Oenotheras  to  be  reached. 

I 


I 


2 


BOTANICAL  GAZETTE 


[JULY 


The  cytological  studies  presented  here  will  be  confined  chiefly  to 
the  phenomena  of  synapsis  and  reduction  in  the  pollen  mother  cell. 
Various  forms  have  been  studied,  a  complete  series  of  stages  being 
obtained  ir^  some  forms  and  a  partial  series  or  only  a  few  stages  being 
examined  in  j6thei,;s:.  /  riie  forms  investigated  include  (i)  O.  rubri- 
nerviSj  (/>)  (9;.  Lamqrckianay(^)  O.  gigas^  (4)  O.  nanella^  (5)  O.  biennis 
cruciate,  ai^ariety  of  the  Eufoijcan  O.  biennis,  (6)  both  O.  lata  (see  12) 
and  O.  Lamorekiana  from  the  Fj  of  O.  lataX  O.  Lamar ckiana,  and  (7) 
plants  resembling  Q'.  ‘pgas,  from  the  Fj  of  O.  lataXO.  gigas.  Pre¬ 
liminary  reports  have  already  been  made  upon  some  of  these  studies, 
in  various  connections  (ii,  12,  13,  14,  15).  Reference  will  be  made 
to  some  of  these  results  later. 

The  material  from  each  individual  was  collected  separately  in 
nearly  all  cases,  in  order  to  observe  possible  individual  differences  in 
the  same  race^  either  in  the  number  of  chromosomes  or  in  other 
cytological  features.  I  am  indebted  to  Mr.  C.  H.  Shattuck  for 
making  a  number  of  these  collections.  The  material  for  the  study  of 
O.  rubrinervis  was  obtained  from  a  number  of  individuals  grown  in 
two  different  seasons  and  representing  several  strains  derived  from  the 
same  original  pedigree.  Sections  were  cut  from  six  of  these,  and  it 
may  be  stated  here  that  in  O.  rubrinervis  no  individual  differences 
were  discovered,  either  in  the  number  of  chromosomes,  which  was 
14  in  all  cases,  or  in  any  other  features.  In  some  of  the  other  mutants, 
also,  a  number  of  individuals  were  examined.  It  was  found  necessary 
to  reserve  the  account  of  O.  gigas,  which  presents  several  features  of 
special  interest,  for  a  separate  paper.  A  preliminary  report  on  this 
form  has  already  been  made  (14,  15). 

For  various  reasons,  O.  rubrinervis  was  chosen  as  the  most  favor¬ 
able  form  for  a  thorough  study  of  synapsis  and  reduction.  The 
nuclei  and  chromosomes  of  Oenothera  are  small,  and  for  this  reason 
the  selection  of  the  most  favorable  type  for  study  is  a  matter  of  some 
importance.  In  O.  rubrinervis  the  pollen  mother  cells,  although 
they  vary  much  in  size,  are  usually  considerably  larger  than  in 
O.  Lamarckiana,  the  nuclei  being  also  proportionately  larger.  The 
reason  for  this  will  be  explained  later.  The  chromosome  number 
being  low  in  most  of  the  forms  (2^  =  14,  x=j),  they  can  be  counted 
without  any  difficulty,  notwithstanding  their  small  size.  Another 


1908] 


GATES— REDUCTION  IN  OENOTHERA 


3 


notable  advantage  in  comparing  this  with  other  studies  in  reduction 
is  in  the  shape  of  the  chromosomes,  which  are  globular  or  somewhat 
oblong  or  cylindrical  in  most  stages  of  mitosis,  and  are  never  greatly 
elongated  or  looped.  For  this  reason  it  is  a  comparatively  easy 
matter  to  obtain  accurate  counts  of  the  chromosomes  in  the  pollen 
mother  cells  of  any  of  the  forms.  This  shape  is  also  found  to  be  very 
advantageous  in  a  study  of  the  events  of  reduction  following  synapsis. 
The  appearances  are  clear  and  easily  interpreted,  in  striking  contrast 
to  the  forms  with  long  twisted  chromosomes,  such  as  have  been  made 
the  basis  of  many  of  the  studies  on  reduction  in  plants. 

On  the  other  hand,  the  somatic  nuclei  and  chromosomes  are  very 
much  smaller,  and  in  metaphase  the  latter  are  elongated  and  looped, 
making  it  impossible  to  count  them  with  the  same  degree  of  accuracy. 
Some  of  these  appearances  have  already  been  described  elsewhere 
(12,  p.  19).  Thus  while  it  was  found  that  the  chromosomes  could 
be  counted  almost  equally  well  in  pollen  mother  cells  of  all  the 
forms  studied,  O.  ruhrinervis  was  found  to  be  especially  favorable 
for  the  investigation  of  reduction  phenomena,  especially  the  events 
of  synapsis  and  the  prophases  of  the  heterotypic  mitosis.  The 
account  given  here  will  refer  throughout  to  O.  ruhrinervis,  with 
occasional  comparisons  with  other  forms.  Later  papers  will  take 
up  these  other  forms  in  detail,  in  so  far  as  this  is  necessary  after  the 
account  presented  here.  Special  attention  will  be  given  at  that  time, 
in  particular,  to  the  later  stages,  beginning  with  the  telophase  of 
the  heterotypic  mitosis,  and  also  to  the  interesting  conditions  in  some 
of  the  hybrids.  The  detailed  account  in  O.  ruhrinervis  will  not  be 
carried  farther  than  the  metaphase  of  the  heterotypic  mitosis,  at 
which  time  the  essential  events  have  already  taken  place. 

Methods 

The  usual  methods  of  cytological  technique  were  employed, 
various  chrom-acetic  and  chrom-osmo-acetic  solutions  being  tried 
until  satisfactory  fixation  was  obtained.  The  thickness  of  the  sec¬ 
tions-  varied  from  4  to  10  /-t.  The  latter  thickness  was  found  most 
favorable  for  counting  chromosomes,  because  it  is  somewhat  greater 
than  the  diameter  of  the  nuclei,  many  of  which  in  such  sections  were 
therefore  uncut.  It  is  possible  to  determine  easily  whether  a  nucleus 
has  been  cut  by  the  knife  by  examining  it  in  low  and  high  focus.  The 


4 


BOTANICAL  GAZETTE 


[JULY 


chromosomes  in  such  uncut  nuclei  can  then  be  counted  with  absolute 
accuracy,  either  in  the  prophase  of  the  heterotypic  mitosis  before  the 
disappearance  of  the  nuclear  membrane,  or  in  the  telophase  after  the 
walls  of  the  daughter  nuclei  are  formed.  In  nearly  every  individual 
examined,  large  numbers  of  such  cases,  all  yielding  the  same  result, 
were  counted  before  the  number  was  finally  determined  upon.  The 
chromosomes  could  also  be  counted  in  certain  positions  on  the 
spindle,  particularly  in  anaphases,  but  in  metaphase  they  were  usually 
too  closely  grouped  to  allow  of  satisfactory  counting. 

In  the  second  division,  particularly  in  the  forms  having  seven 
chromosomes  as  the  gametophytic  number,  the  chromosomes  could 
be  counted  with  certainty  in  almost  any  stage  of  mitosis.  The  thinner 
sections  were  used  chiefly  in  the  study  of  spirem  and  synapsis  stages, 
although  here  also  the  comparatively  short  length  of  the  thickened 
spirem  frequently  made  it  advantageous  to  study  uncut  nuclei  in 
which  the  spirem  could  be  followed  throughout  its  length. 

Of  the  various  stains  Heidenhain’s  iron-hematoxylin  was  found 
to  be  superior  for  chromosome  counting  and  for  clear  differentiation 
of  chromatic  structures  in  nearly  all  stages  of  synapsis  and  reduction, 
safranin-gentian  being  used  occasionally  for  comparison  or  for 
differentiating  particular  cytoplasmic  structures.  Orange  G  was 
also  used  with  the  iron-alum  stain  for  bringing  out  clearly  certain 
special  features,  particularly  the  protoplasmic  connections  between 
mother  cells,  which  will  be  described  later. 

Description 

EARLY  STAGES 

Some  of  the  very  early  stages  of  the  anthers,  previous  to  the  forma¬ 
tion  of  mother  cells,  have  been  studied  particularly  with  the  purpose 
of  tracing  the  origin  of  the  bodies  which  were  called  heterochromo¬ 
somes  in  my  first  paper.  The  provisional  use  of  the  name  was 
based  on  the  very  close  resemblance  of  these  bodies  to  the  chromo¬ 
somes  in  appearance,  and  their  frequent  presence  close  by,  or  in  some 
cases  apparently  attached  to,  the  heterotypic  spindle.  They  were 
not  stated  to  pass  undivided  into  one  of  the  daughter  nuclei,  as 
misquoted  by  Tischler  (32),  but  to  remain  outside  in  the  cytoplasm 
where  they  gradually  disappear.  The  study  of  their  early  history 


1908] 


GATES— REDUCTION  IN  OENOTHERA 


5 


shows  that  no  line  of  distinction  can  be  drawn  between  them  and  the 
large  body  readily  recognized  as  the  nucleolus.  They  are  then 
smaller  nucleoli,  not  differing  essentially  in  origin  from  the  single 
larger  body  which  is  almost  constantly  present  in  the  mother  cell 
during  synapsis  and  prophase,  but  diverging  from  the  latter  some¬ 
what  in  their  later  history. 

In  the  earliest  stages  studied,  the  young  meristematic  cells  of  the 
anther  primordia  are  very  small  {figs.  /,  2),  and  the  tissues  are  wholly 
undifferentiated,  except  the  epidermal  layer.  Usually  several  smaller 
nucleoli  are  present  in  each  nucleus  of  the  meristematic  cells,  in  addi¬ 
tion  to  the  larger  nucleolus.  Compared  with  the  cells  of  the  anther 
wall  when  they  are  no  longer  meristematic,  the  smaller  nucleoli  of 
the  former  are  about  the  size  of  the  nucleoli  of  the  latter,  which  are 
approximately  equal  in  size.  There  is  nothing  in  the  latter  corre¬ 
sponding  to  the  larger  nucleolus  of  the  meristematic  cells.  Probably 
afterward  one  of  these  nucleoli  enlarges  as  the  cell  increases  in  size, 
or  it  is  possible  that  the  nuclei  of  meristematic  cells  are  always  derived 
from  previous  ones  which  already  possess  a  large  nucleolus. 

‘  Chromatic  staining  bodies  are  also  found  closely  appressed  to  the 
nuclear  membrane  in  all  the  meristematic  cells  {figs,  i,  2).  This 
tendency  for  chromatic  material  to  accumulate  on  the  nuclear  walls 
gives  these  nuclei  a  characteristic  appearance.  These  bodies  often 
appear  like  a  thickening  of  the  membrane  itself. 

At  the  next  stage  studied  many  cell  divisions  have  taken  place, 
and  the  sporogenous,  tapetal,  and  wall  tissues  have  been  differentiated. 
The  sporogenous  cells  have  increased  enormously  in  size,  and  form 
a  single  row  in  longitudinal  section  down  the  center  of  the  anther,  the 
walls  of 'these  cells  being  especially  thickened  and  distinct  {fg.  fi). 
The  cells  of  the  surrounding  tapetal  layer  have  also  increased  greatly 
in  size  and  are  distinctly  marked  off  from  the  sporogenous  row.  In 
the  sporogenous  cells  the  nuclei  {fg.  4),  though  much  increased  in 
size,  have  not  increased  in  proportion  to  the  cytoplasm.  The  large 
nucleolus,  much  larger  than  in  the  earlier  stage,  is  now  a  conspicuous 
object  in  the  nucleus.  Smaller  nucleolar  bodies  are  almost  invariably 
present,  but  masses  are  no  longer  found  attached  to  the  nuclear 
membrane.  (The  characteristic  masses,  however,  may  remain  for 
some  time  attached  to  the  nuclear  walls  of  the  tapetal  cells) . 


6 


BOTANICAL  GAZETTE 


[JULY 


Figs,  j-io  are  from  drawings  of  other  nuclei  at  this  stage  of  develop¬ 
ment.  In  the  majority  of  cases  one  or  two  smaller  nucleoli  occur 
in  addition  to  a  single  large  one,  but  rarely  {fig.  6)  two  large  nucleoli 
of  equal  size  may  be  found ;  and  very  frequently  the  number  of  small 
bodies,  of  equal  or  unequal  size,  may- be  greater,  reaching  as  many  as 
five  or  six.  Figs.  5,  y,  8,  g  show  these  in  various  stages  of  fusion  with 
each  other  and  with  the  large  nucleolus.^  They  are  thus  not  in  any 
sense  autonomous  bodies.  It  appears  that  usually  these  fusions 
take  place  until  only  one  large  nucleolus  and  one  or  two  smaller  ones 
are  present  during  synapsis  and  diakinesis.  But  occasionally  the 
fusions  do  not  take  place,  and  several  of  these  bodies  may  then  be 
present  in  the  later  stages.  The  number  of  these  nucleoli  finally 
present  depends,  then,  largely  upon  the  amount  of  fusion  which  has 
previously  taken  place  between  them.  In  the  later  stages  one  large 
nucleolus  is  almost  invariably  present  and  usually  a  smaller  one 
bearing  a  certain  proportion  to  the  larger  in  size,  though  the  latter 
may  vary  in  size  and  number  as  already  stated.  There  is  usually 
a  clear  area  around  the  large  nucleolus,  as  in  the  earlier  stage,  and 
threads  of  the  reticulum  may  or  may  not  cross  this  and  appear  to  be 
attached  to  the  nucleolus  {fig.  4).  The  reticulum  of  the  cytoplasm 
usually  stains  rather  more  deeply  at  this  time  than  that  of  the  nucleus. 
It  may  as  well  be  stated  at  this  time  that  in  the  resting  nuclei  of  the 
pollen  tetrad  and  in  the  nuclei  of  the  nearly  mature  pollen  grains  of 
Oenothera  one  finds  {fig.  ii)  the  same  condition  of  the  nucleoli  as 
in  the  mother  cells,  namely,  usually  one  large  and  one  small  nucleolus 
bearing  a  rather  definite  size  relation  to  each  other,  with  sometimes 
additional  small  ones. 

The  sporogenous  rows  are  differentiated  from  the  tapetum  by 
the  greater  growth  of  the  cells,  nuclei,  and  nucleoli  of  the  former. 
At  the  same  time  they  are  distinctly  marked  off  by  the  formation  of  a 
continuous  thickened  wall  between  tapetum  and  archesporium 
{fig.  j).  It  is  obvious  that  as  the  cells  and  nuclei  increase  in  size, 
the  nucleolus  grows  also.  Up  to  the  time  of  synapsis  the  mother  cells 
usually  form  a  compact  tissue,  but  about  this  time  the  cells  begin  to 

I  Miss  Nichols  (21)  figures  what  are  in  all  probability  stages  of  fusion  of  large 
and  small  nucleoli  in  Sarracenia  pollen  mother  cells,  but  interprets  them  as  a  budding- 
off  of  small  bodies  from  the  nucleolus.  The  budlike  attachments  to  the  nucleolus, 
frequently  observed  by  other  authors  are  doubtless  to  be  explained  in  like  manner. 


1908] 


GATES— REDUCTION  IN  OENOTHERA 


7 


break  apart  at  the  corners  where  they  meet,  and  before  diakinesis  is 
reached  they  are  completely  rounded  off  and  independent,  or  they 
frequently  remain  connected  with  other  mother  cells  only  at  the  ends. 
In  the  meantime  the  cavity  of  the  loculus  grows  rapidly,  so  that  the 
mother  cells,  in  normal  development,  usually  lie  loose  in  the  cavity. 

The  events  of  synapsis  and  reduction  usually  go  forward  simul¬ 
taneously  throughout  a  flower,  with  comparatively  little  variation  in 
the  different  parts  of  the  same  loculus  or  in  the  different  anthers  of  a 
flower.  In  one  flower,  however,  wide  variation  was  found  in  different 
anthers,  but  comparative  constancy  in  the  loculus.  One  anther  of 
this  flower  was  in  synapsis,  another  in  diakinesis,  another  in  meta¬ 
phase  of  the  heterotypic  mitosis,  and  in  still  another  some  of  the 
mother  cells  had  completed  the  second  mitosis.  No  abnormalities 
in  the  cytological  condition  of  this  flower  were  observed. 

SYNAPSIS 

After  the  stage  described  in  fig.  4,  the  nucleus  increases  greatly  in 
size,  but  without  an  appreciable  increase  in  the  size  of  the  cell.  The 
single  row  of  sporogenous  cells  divides,  so  that  there  are  usually  two 
rows  of  pollen  mother  cells.  Occasionally  three  or  more  mother 
cells  are  found  in  the  cross-section  of  a  loculus.  In  general  there  are 
fewer  divisions  than  in  the  other  forms,  and  this  is  at  least  one  of  the 
reasons  why  the  mother  cells  are  on  the  average  larger  than,  for 
example,  in  O.  Lamarckiana. 

The  resting  nucleus  of  the  pollen  mother  cell  increases  in  size  and 
begins  to  show  signs  of  approaching  synapsis.  Figs.  12,  ij,  14 
show  stages  in  the  beginning  of  this  process.  A  number  of  these 
stages  were  found — although  they  are  uncommon — ^in  the  same 
sections  with  regular  synapsis  stages.  In  some  cases  they  occurred 
side  by  side  with  mother  cells  in  which  the  synaptic  knot  had  already 
been  formed.  A  complete  series  of  stages  may  be  found  in  the  same 
section,  from  the  beginning  of  contraction  to  the  formation  of  a  close 
synaptic  ball.  The  cytoplasm  in  these  cells  shows  no  contraction 
whatever,  but  is  perfectly  fixed.  For  this  and  other  reasons  there 
can  be  no  doubt  that  this  is  a  real  contraction  stage,  leading  to  synap¬ 
sis,  and  not  a  result  of  imperfect  fixation,  as  one  might  judge  on  first 
examination. 


8 


BOTANICAL  GAZETTE 


[JULY 


That  these  nuclei  are  going  into  synapsis  and  not  coming  out  is 
shown  by  several  features:  (i)  the  extremely  delicate  character  of  the 
threads,  like  those  of  the  resting  nucleus;  (2)  the  fact  that  the 
periphery  of  the  reticulum  as  it  contracts  frequently  preserves  perfectly 
the  curved  outline  of  the  nuclear  wall  {jig.  12) ;  (3)  immediately  after 
synapsis  the  thread  is  somewhat  shorter  and  thicker  than  previously 
and  appears  to  be  continuous,  while  in  the  earlier  contraction  stages 
we  still  have  the  appearance  of  a  reticulum  {jig.  ij).^  As  the  con¬ 
traction  progresses,  the  threads  are  gradually  rearranged  from  an 
anastomosing  reticulum  to  a  very  long  and  continuous  delicate  thread. 
The  exact  manner  of  this  rearrangement  could  not  be  observed,  but 
one  finds  many  transitions  {jig.  14)  from  the  anastomosing  reticulum 
of  the  resting  nucleus  to  the  closely  coiled  and  apparently  continuous 
spirem  of  the  synaptic  knot  {jig.  15).  The  contraction  may  take 
place  from  one  side  of  the  nucleus,  leaving  the  reticulum  attached  for  a 
time  to  the  nuclear  membrane  at  one  point  {jig.  ij),  or  it  may  take 
place  simultaneously  from  all  sides  {fig.  12).  A  few  threads  of  the 
reticulum  usually  remain  attached  for  a  time  to  the  nuclear  membrane 
while  the  contraction  is  going  on.  These  are  drawn  in  finally  as  the 
synaptic  ball  becomes  more  compact. 

The  small  number  of  these  intermediate  stages  found  indicates 
that  they  are  passed  through  rather  rapidly,  the  frequency  of  the  occur¬ 
rence  of  synapsis  stages  indicating,  on  the  other  hand,  that  this  con¬ 
dition  is  of  considerable  duration. 

No  indication  of  a  doubling  or  pairing  of  the  threads  during  these 
intermediate  contraction  stages  could  be  observed,  though  they  were 
carefully  searched  for.  Moreover,  in  the  earliest  stages  of  the  synaptic 
ball  the  thread  appears  to  be  as  thin  and  delicate  as  in  the  reticulum, 
which  does  not  favor  the  view  that  a  pairing  has  taken  place.  The 
evidence,  then,  £0  far  as  it  goes,  is  decidedly  not  in  favor  of  a  pairing. 

During  these  stages  the  nuclear  membrane  is  often  indistinct, 
making  it  difficult  to  define  accurately  the  limits  of  the  nucleus.  The 

2  This  explanation  assumes,  of  course,  that  the  synapsis  stages  themselves  are 
normal  and  not  due  to  artifact,  as  I  presume  all  cytologists  will  now  agree,  although 
ScHAFFNER  (29)  apparently  still  entertains  some  doubt  on  the  subject.  The  regularly 
coiled  arrangement  of  the  thread  in  the  synaptic  ball  appears  to  me  to  be  one  of  the 
best  arguments  against  this  stage  being  an  artifact.  Evidently  a  rearrangement  of 
the  threads  is  going  on  as  contraction  proceeds. 


1908] 


GATES— REDUCTION  IN  OENOTHERA 


9 


same  condition  is  observed  during  synapsis,  which  is  found  in  the 
same  sections.  In  places  the  nuclear  membrane  has  either  disap¬ 
peared  or  is  too  delicate  to  be  observed.  The  cytoplasm,  however, 
retains  the  original  outline  of  the  nucleus.  Mottier  (20)  has 
apparently  observed  similar  conditions  of  the  nuclear  membrane  at 
this  time.  In  some  cases  it  is  ruptured  and  a  portion  of  it  is  actually 
carried  inward  with  the  nuclear  reticulum  at  the  beginning  of  the 
contraction  {-jig.  12).  One  is  tempted  to  explain  this  as  an  artifact; 
but  that  this  is  not  the  explanation  is  shown  by  the  considerations 
already  mentioned.  The  explanation  appears  to  be  that  as  contrac¬ 
tion  proceeds  a  portion  of  the  nuclear  membrane  may  be  torn  away 
and  carried  inward  attached  to  the  threads.  Frequently  in  these 
stages  one  finds  the  nuclear  membrane  present  on  one  side  of  the 
nucleus  but  invisible  elsewhere.  This  is  the  case  in  jig.  12,  although 
the  membrane  was  drawn  as  though  complete.  Observations  of  other 
nuclei  bear  out  this  interpretation,  the  nuclear  membrane  being  clearly 
visible  in  some  cases  attached  to  portions  of  the  reticulum  which  have 
contracted  far  away  from  the  original  position  of  the  nuclear  wall.  In 
the  late  prophase,  when  the  definitive  chromosomes  are  formed,  a 
distinct  and  perfect  nuclear  membrane  is  invariably  present,  so  it 
would  appear  that  in  such  cases  as  those  just  described  a  new  mem¬ 
brane  is  afterward  formed. 

Mention  must  now  be  made  of  the  chromatic  staining  material  of 
the  nucleus  during  these  stages.  The  nucleolus  is  frequently,  though 
not  always,  included  within  the  synaptic  knot.  There  is  a  tendency 
for  other  dark-staining  bodies  to  accumulate  near  the  periphery  of 
the  nucleus  {jigs.  12,  14) ;  as  contraction  proceeds  these  are  swept  in 
by  the  reticulum.  The  exact  relation  they  bear  to  the  threads  is  not 
known.  In  some  cases  they  appear,  in  the  later  stages  of  contraction 
at  least,  to  form  a  part  of  the  threads  themselves,  in  other  cases  they 
appear  to  be  merely  inclusions  in  its  coils.  These  bodies  show  no 
constancy  in  number,  size,  or  shape.  As  the  spirem  takes  on  the  ap¬ 
pearance  of  the  synaptic  knot,  they  are  still  found  in  its  meshes,  and 
portions  of  the  thread  itself  may  also  stain  darkly,  suggesting  a 
solution  of  a  part  of  their  substance  and  its  transfer  into  the  thread. 
Even  when  the  greater  part  of  the  spirem  is  completely  decolorized 
certain  portions  of  it  retain  the  stain.  This  appears  to  be  partly 


lO 


BOTANICAL  GAZETTE 


[JULY 


due  to  the  denser  aggregation  of  the  spirem  in  these  regions,  but 
in  some  cases  it  is  evidently  due  to  the  presence  of  bodies  which  retain 
the  stain  and  appear  to  be  giving  up  the  stainable  part  of  their  sub¬ 
stance  to  the  spirem.  These  bodies  are  evidently  not  the  prochromo¬ 
somes  found  by  Overton  (22)  in  certain  dicotyledons,  nor  are  they 
the  gamosomes  of  Strasburger  (30,  31). 

Just  the  relation  these  bodies  sustain  to  the  spirem  is  not  easy  to 
determine.  From  figs.  12  and  14  it  is  evident  that  they  are  at  first 
small  “nucleoli”  caught  in  the  contracting  reticulum,  but  quite 
independent  of  it.  Later  they  appear  to  give  up  a  portion  at  least  of 
their  material  to  the  spirem,  finally  disappearing  as  independent 
bodies.  Usually,  however,  at  least  one  of  these  bodies  remains  inde¬ 
pendent,  and  appears  in  synapsis  and  diakinesis  as  a  small  nucleolus 
bearing  a  definite  relation  to  the  size  of  the  large  nucleolus,  being 
about  the  size  of  a  chromosome.  These  bodies  are  usually  free  in 
the  nuclear  cavity  {fig.  ij).  A  certain  depth  of  stain  is  required  for 
demonstrating  them  during  synapsis,  for  they  usually  decolorize  more 
quickly  than  the  large  nucleolus.  With  a  favorable  stain  they  are 
found  to  be  of  strikingly  uniform  occurrence  at  this  time.  A  plasma 
stain  such  as  orange  G  may  be  used  with  advantage  to  demonstrate 
their  presence.  The  uniformity  in  their  occurrence  is  so  great  that 
for  some  time  they  were  thought  to  be  constant  in  size  and  number. 
With  the  demonstration  of  their  inconstancy  and  their  origin  we  have 
chosen  to  call  them  merely  small  nucleoli,  as  there  appears  to  be  no 
sufficient  reason  for  another  name.  The  (large)  nucleolus  disappears 
with  great  promptness  immediately  after  the  nuclear  membrane  breaks 
down,  only  persisting  for  a  time  in  a  few  rare  instances.  In  no  case 
has  fragmentation  of  the  nucleolus,  previous  to  its  disappearance,  been 
observed,  although  the  presence  of  deeply  staining  globular  bodies 
occasionally  found  near  the  periphery  of  the  cytoplasm  might  be 
accounted  for  in  this  way.  The  mass  of  the  latter,  however,  is  some-  • 
times  greater  than  that  of  the  nucleoli.  The  smaller  nucleoli  persist 
and  are  frequently  found  close  by  the  heterotypic  spindle.  They 
may  also  be  found  on  the  homotypic  spindle  {fig.  41).  Apparently 
they  never  reenter  a  nucleus,  but  remain  in  the  cytoplasm  until  they 
finally  disappear.  These  bodies  have  been  found  showing  the  same 
behavior  in  all  the  forms  studied. 


% 


4 


1908] 


GATES— REDUCTION  IN  OENOTHERA 


II 


POST-SYNAPTIC  STAGES 

Synapsis  lasts  for  a  comparatively  long  time,  as  shown  by  the  fre¬ 
quency  of  its  occurrence  in  the  material  sectioned.  During  this  time 
the  spirem  shortens  and  thickens  and  then  begins  to  arrange  itself 
more  loosely  in  the  nuclear  cavity.  This  shortening  and  thickening 
is  progressive  {jigs.  16-18)  and  apparently  continues  for  some  time. 
During  these  stages  the  thickness  of  the  spirem  may  be  nearly  uniform 
throughout,  or  it  may  vary  greatly,  giving  a  moniliform  appearance,  or 
the  spirem  may  appear  irregularly  constricted  at  varying  intervals. 
In  other  cases,  with  a  certain  depth  of  stain  it  is  seen  to  be  composed 
of  lighter  and  darker  areas  more  or  less  regularly  alternating.  Por¬ 
tions  of  the  thread  may  appear  homogeneous  or  may  show  the  lighter 
and  darker  areas,  according  to  the  depth  of  stain  {jig.  if).  In  more 
deeply  stained  nuclei,  such  as  jig.  16,  the  thread  appears  homogeneous 
throughout.  These  darker  areas  are  the  chromatin  discs  or  chromo- 
meres  of  various  authors;  and  they  give  the  thread  a  very  character¬ 
istic  appearance.  During  this  well-defined  stage  the  greatly  thickened 
spirem  is  loosely  distributed  in  the  nuclear  cavity.  Deeply  staining 
bodies  still  appear  attached  to  or  enmeshed  in  the  coils  of  the  thread. 

At  this  time  one^  finds  undoubted  indications  of  parallel  threads. 
When  represented  by  camera  drawings  in  one  plane  the  evidence  for 
this  is  not  so  satisfactory  as  in  the  original  preparation,  but  there  is  no 
doubt  of  their  occurrence.  As  already  stated,  in  the  earlier  stages 
previous  to  and  during  synapsis,  parallel  threads  could  not  be  observed, 
and  it  has  not  been  determined  whether  they  were  really  absent  or 
whether  the  failure  to  observe  them  was  due  to  their  extreme  delicacy. 
Hence  it  cannot  now  be  stated  whether  they  have  arisen  through  an 
approximation  of  spirems  at  an  earlier  period,  or  through  a  split  in  the 
single  continuous  spirem.  This  matter  will  be  discussed  later. 

Following  this  stage  a  second  well-marked  contraction  takes  place 
{jigs.  18,  20,  21),  apparently  quite  as  typical  and  constant  in  its  occur¬ 
rence  as  the  first  contraction  stage,  which  is  ordinarily  identified  as 
synapsis.  This  contraction  is  of  much  shorter  duration,  however, 
and  entirely  different  in  appearance,  owing  to  changes  which  the 
thread  has  undergone  since  synapsis,  resulting  in  a  great  amount  of 
shortening  and  thickening  of  the  spirem.  Mottier  (20)  has  recog¬ 
nized  this  second  contraction  stage  in  Podophyllum,  Lilium,  and 


12 


BOTANICAL  GAZETTE 


[JULY 


Tradescantia,  though  he  formerly  thought  it  resulted  from  bad  fixa¬ 
tion  ;  and  it  appears  to  have  been  observed  also  by  Farmer  and  Shove 
(lo).  Mottier  states  that  in  these  forms  there  is  but  little  shorten¬ 
ing  of  the  spirem  between  synapsis  and  segmentation  into  chromo¬ 
somes.  In  Oenothera,  on  the  contrary,  as  is  evident  from  a 
comparison  of  'jigs.  15  or  id  with  22,  a  very  considerable  amount  of 
shortening  as  well  as  thickening  of  the  spirem  takes  place  during  this 
interval.  During  the  second  contraction  the  paired  threads  apparently 
fuse,  and  further  shortening  of  the  (from  now  single)  thread  results  in 
an  enormous  amount  of  thickening  of  the  spirem,  so  that  when  it  uncoils 
from  this  second  contraction  it  has  approximately  the  thickness  of  a 
chromosome  and  exhibits  only  a  few  loops.  It  can  then  frequently 
be  traced  through  nearly  its  whole  length.  At  this  time  there  is  a 
great  amount  of  variation  in  the  thickness  of  different  parts  of  the 
spirem,  as  seen  in  jigs.  22  and  2j.  Fig.  ig  is  a  portion  of  the  spirem 
at  this  period,  drawn  with  a  higher  magnification.  It  shows  the 
chromatic  bodies,  which  vary  in  size,  imbedded  in  the  linin  substra¬ 
tum.  As  to  how  far  two  different  substances  are  represented,  I  am 
at  present  unprepared  to  say. 

DIAKINESIS 

The  single  thick  thread  now  segments  transversely  into  14  chromo¬ 
somes,  the  sporophyte  or  2X  number.  At  this  time  there  is  no  indica¬ 
tion  whatever  of  a  longitudinal  split  in  the  thread.-  Even  when  greatly 
washed  out,  the  material  of  the  chromosomes  appears  perfectly 
homogeneous,  or  if  a  granular  structure  is  observable  there  is  in  its 
arrangement  no  indication  of  the  previous  split.  At  the  time  of  this 
second  contraction  a  pair  of  chromosomes  is  frequently  observed 
separated  from  the  spirem  and  apparently  always  lying  with  their 
long  axes  parallel  and  connected  at  one  end  {figs.  20,  22).  This 
condition  occurs  very  commonly,  although  in  other  cases  the  spirem 
is  continuous  throughout  (jig.  21).  In  no  case  has  more  than  one 
pair  of  chromosomes  been  observed  to  be  thus  precociously  cut  off  in 
O.  ruhrinervis ,  though  two  such  pairs  have  been  observed  in  O.  lata 
(see  1 1 ,  jig.  ig) .  In  no  case  has  a  single  chromosome  been  observed 
to  be  cut  off  in  this  manner,  and  apparently  they  are  invariably  cut 
off  in  pairs,  that  is,  bivalvent  chromosomes  are  detached. 


1908] 


GATES— REDUCTION  IN  OENOTHERA 


13 


What  significance  this  early  separation  of  chromosome  pairs  may 
have  is  not  known,  but  it  appears  that  the  later  history  of  these  pairs 
on  the  spindle  can  be  traced.  In  the  paper  just  cited  (ii),  the 
writer  wrongly  identified  them  with  the  smaller  nucleoli  which  persist 
by  the  heterotypic  spindle.  These  chromosome  pairs  are  frequently 
so  closely  approximated  at  the  end  opposite  the  end  of  actual  connec¬ 
tion  as  to  give  the  appearance  of  a  ring.  It  was  thought  that  these 
rings  by  condensation  (which  actually  takes  place)  were  reduced  to 
the  size  of  these  nucleolar  bodies.  The  latter  had  the  size  and  shape 
of  chromosomes,  and  with  a  certain  depth  of  stain  invariably  appeared 
hollow.  These  pairs  are  not  condensed  to  rings,  however,  but  to 
chromosome  pairs  of  the  ordinary  Oenothera  type. 

The  spirem  at  this  time  varies  greatly  in  thickness  in  different 
parts,  exhibiting  constrictions  and  dilatations  which  indicate  more 
or  less  clearly  where  segmentation  into  chromosomes  will  take  place. 
This  segmentation  may  happen  while  the  spirem  is  still  in  the  con¬ 
tracted  condition  {jig.  25) ,  or  after  it  has  again  uncoiled  and  distrib¬ 
uted  itself  in  the  nuclear  cavity  {jigs.  24,  26,  28),  or  before  this 
uncoiling  is  completed.  The  segmentation  appears  to  be  in  some 
cases  nearly  simultaneous  {jig.  24) ;  in  other  cases  the  segmentation  is 
successive,  as  in  jig.  2j,  where  the  spirem  is  clearly  divided  into  three 
portions  and  the  constrictions  for  the  formation  of  the  chromosomes 
are  so  far  advanced  that  the  number  of  chromosomes  to  be  formed 
by  each  segment  can  already  be  foretold  with  practical  certainty. 
The  segmentation  at  this  time  is  into  14  chromosomes,  the  sporophyte 
number.  •  A  large  number  of  counts  made  at  this  time  demonstrate 
the  absolute  constancy  of  this  number  in  all  the  individuals  of  O. 
ruhrinervis  examined.  It  is  possible,  however,  that  individuals  of  this 
race  may  be  found  whose  chromosome  number  differs  from  this 
number  by  one.  This  matter  will  be  discussed  later. 

In  every  single  case  where  the  count  could  be  determined  with 
certainty  it  was  shown  to  be  14.  These  counts  were  all  made  from 
sections  10  //-  thick,  and  from  nuclei  which  were  uncut  by  the  knife. 
The  less  numerous  counts  made  in  the  multipolar  stage  of  the  hetero¬ 
typic  spindle  gave  invariably  the  same  number.  In  this  case  all  in  a 
given  cell  were  obtained  by  examining  the  adjacent  sections.  In  all, 
hundreds  of  counts  were  made.  In  such  nuclei  as  jigs.  26,  2g,  jo,  ji 


14 


BOTANICAL  GAZETTE 


[JULY 


there  can  be  no  possible  doubt  of  the  number  of  chromosomes 
present. 

As  already  shown  {fig.  20),  one  or  in  some  cases  more  pairs  of 
chromosomes  may  be  cut  off  from  the  spirem  before  it  undergoes 
segmentation,  and  frequently  while  it  is  still  in  the  second  contraction 
period.  The  exact  method  of  origin  of  these  pairs  has  not  been 
observed,  but  they  invariably,  so  far  as  observed,  lie  with  their  long 
axes  parallel  and  connected  at  one  end,  from  which  it  would  appear 
that  they  were  successive  chromosomes  on  the  spirem.  In  later 
stages,  when  the  spirem  has  constricted  into  a  chain  of  chromosomes 
arranged  near  the  periphery  of  the  nucleus,  one  or  more  pairs  of 
chromosomes  are  found  separated  from  the  rest.  Some  of  these  have 
doubtless  had  the  origin  shown  in  fig.  20.  Others  appear  to  have 
originated  later,  as  indicated  in  some  of  the  figures,  by  successive 
chromosomes  on  the  chain  swinging  around  parallel  to  each  other  and 
thus  pairing.  Usually  in  diakinesis  one  or  two  such  pairs  are  found, 
though  occasionally  there  is  no  evidence  of  pairing.  The  highest 
number  of  pairs  observed  at  this  stage  was  five,  with  indications  of 
pairing  among  the  others  {fig.  2g) ;  which  is  unusual.  Later,  in  the 
multipolar  spindle  stage  two  distinct  pairs  are  usually  found  in  vary¬ 
ing  stages  of  conjugation  {figs,  jy,  jd).  A  single  case  was  observed 
{fg.  J7)  in  which  the  fourteen  chromosomes  were  all  paired. 

As  the  figures  indicate,  constriction  of  the  spirem  at  regular  inter¬ 
vals  proceeds  progressively  until  a  chain  of  chromosomes  is  formed. 
When  this  has  taken  place,  the  chromosomes  are  at  first  several 
times  longer  than  broad,  and  their  margins  have  a  very  irregular, 
sinuous  outline,  like  that  of  the  spirem  just  previous  to  segmentation. 
They  are  not  so  long,  however,  that  they  can  be  twisted  and  looped 
in  the  confusing  manner  of  many  heterotypic  chromosomes  of  plants. 
This  is  very  gratifying  in  the  study  of  these  stages,  since  it  permits 
a  clearness  of  interpretation  which  would  otherwise  be  unattainable. 
Figs.  22  and  show  the  beginning  of  contraction,  which  has  pro¬ 
ceeded  farther  in  fg.  24,  leaving  only  the  so-called  linin  connection 
between  the  chromosomes.  The  constrictions  are  all  equivalent  and 
the  spirem  thus  segments  into  the  sporophyte  number  of  chromosomes 
and  not  into  the  reduced  number  of  chromosome  pairs.  If  successive 
chromosomes  on  the  spirem  are  really  the  members  of  a  pair,  there  is 


1908} 


GATES— REDUCTION  IN  OENOTHERA 


15 


nothing  in  the  manner  of  segmentation  of  the  spirem  to  indicate  this. 
However,  it  is  clear  enough  that  one  chromosome  frequently  swings 
around,  as  already  mentioned,  and  pairs  with  its  neighbor  on  the 
spirem.  We  do  not  really  have,  then,  a  transverse  division  of  chromo¬ 
some  bivalents,  but  a  separation  of  whole  (somatic)  chromosomes. 
Nothing  has  been  found  in  the  earlier  stages  which  would  correspond 
to  the  gamosomes  and  zygosomes  of  Strasburger,  and  even  should  a 
pairing  of  parallel  threads  during  synapsis  occur  (a  possibility  which 
will  be  discussed  later),  the  final  pairing  is  between  chromosome 
bodies  which  were  lying  end  to  end  on  a  single  spirem  thread. 

The  linin  connections  during  diakinesis  appear  to  be  merely  the 
more  finely  drawn  out  portion  of  the  spirem  between  the  chromosomes. 
As  condensation  and  contraction  of  the  chromosomes  progress, 
these  linin  connections  become  longer  and  more  delicate  {jigs,  ji,  jj). 
The  chromosomes  become  more  dense  and  compact,  being  at  first 
oblong-cylindrical  {figs.  24,  26)  and  then  more  nearly  globular  or 
pear-shaped  {fig.  ji).  Certain  chromosomes  sometimes  undergo  this 
contraction  more  quickly  than  others,  as  in  fig.  2g,  and  the  different 
stages  of  this  condensation  may  occasionally  all  be  found  in  the  same 
nucleus.  In  other  cases  the  globular  appearance  is  due  to  the 
position  in  which  certain  chromosomes  happen  to  be  lying  {fig.  J4). 

HETEROTYPIC  MITOSIS 

During  the  prophase  stages  last  outlined  the  cytoplasm  usually  pos¬ 
sesses  a  more  or  less  obscurely  radiate  appearance.  A  felt-work 
of  fibrillae  finally  appears  around  the  nuclear  membrane.  Later 
these  fibrillae  come  to  run  tangentially  to  the  latter,  terminating  in  the 

cytoplasm,  and  by  their  aggregation  in  certain  regions  the  multipolar 

_  • 

spind'e  is  formed.  From  this  stage  the  fibers  are  rearranged  to  form 
the  bipolar  spindle,  passing  through  conditions  in  which  the  spindle 
appears  quadripolar  or  tripolar  in  section.  In  the  meantime  the 
nuclear  membrane  has  dissolved  and  the  chromosomes  are  found 
at  first  in  a  cavity  surrounded  by  fibers  which  preserve  the  outline  of 
the  nuclear  wall.  Later  they  come  in  and  become  attached  to  the 
chromosomes.  Usually  the  large  nucleolus  has  vanished  before  this 
time,  but  occasionally  it  may  still  be  seen  {fig.  J5).  In  fig.  jy  the 
small  nucleolus  is  shown,  which  can  very  frequently  be  seen  at  this 


i6 


BOTANICAL  GAZETTE 


[JULY 


time.  Figs.  j6  and  j/  are  merely  sketches  of  the  spindle  fibers  to 
indicate  their  general  direction.  Fig.  J5  is  an  unusual  case.  A  cone 
of  fibers  appears  to  have  been  formed  on  one  side  only  of  the  nucleus. 
The  fibers  are  coming  in  and  finding  attachment  to  the  chromo¬ 
somes.  The  large  nucleolus  is  still  present,  as  well  as  two  smaller  ones. 

The  most  critical  stages  of  reduction  have  now  been  described  and 
the  remaining  stages  will  be  taken  up  with  less  detail  at  this  time,  but 
will  be  presented  in  full  in  a  later  paper.  The  chromosomes  are  at 
first  irregularly  arranged  on  the  heterotypic  spindle.  As  already 
seen,  during  spindle  formation  many  of  the  chromosomes  are  fre¬ 
quently  separate  and  unpaired.  The  attraction  between  the  chromo¬ 
somes  which  leads  to  pairing  is  evidently  weak,  so  that  it  is  doubtful 
if  any  pairing  takes  place  at  metaphase  between  chromosomes 
which  had  not  previously  paired.  On  the  other  hand,  chromosomes 
which  have  once  paired,  no  matter  how  early,  appear  to  remain 
together  until  their  separation  in  the  metaphase  of  the  heterotypic 
mitosis.  Hence  probably  in  many  cases  the  chromosomes  pass  to  the 
poles  of  the  heterotypic  spindle  without  having  previously  paired  with 
each  other,  that  is,  they  were  merely  lying  loosely  in  the  equatorial 
region  of  the  spindle  in  metaphase,  so  that  it  was  largely  a  matter  of 
chance  which  pole  any  particular  chromosome  went  to.  This  is 
believed  to  be  a  matter  of  prime  importance  in  determining  the  final 
result  of  the  reduction  divisions  in  Oenothera,  and  the  nature  of  the 
distribution  of  chromatin  elements  which  takes  place.  Its  possible 
significance  will  be  pointed  out  in  the  discussion.  Fig.  shows  the 
chromosomes  just  being  drawn  into  the  equatorial  plate  of  the  hetero¬ 
typic  spindle.  In  the  examination  of  thousands  of  spindles  in  about 
this  stage,  one  usually  finds  the  chromosomes  spread  out  in  several 
planes  along  the  long  axis  of  the  spindle.  Of  course  some  of  these  are 
early  anaphase  stages  in  which  the  chromosomes  have  begun  their 
journey  to  the  poles,  but  the  condition  is  seldom  found  where  the 
chromosomes  are  arranged  regularly  in  pairs  on  the  spindle.  The 
daughter  chromosomes  seldom  advance  toward  the  pole  in  a  single 
plane,  as  is  the  case  in  so  many  forms,  but  are  more  or  less  irregularly 
strung  out  along  the  spindle  in  their  passage  to  the  poles.  This  is  in 
striking  contrast  with  their  behavior  in  the  homotypic  mitosis. 

Usually  in  the  early  anaphase  of  the  heterotypic  mitosis  a  longi- 


( 


1908] 


GATES— REDUCTION  IN  OENOTHERA 


17 


tudinal  split  appears  in  the  daughter  chromosomes.  This  split  does 
not  stop  short  of  one  end,  giving  a  V-shaped  body  as>n  niany  plant 
chromosomes,  but  usually  passes  right  jthro'agt.L.JoriTiing  two  inde- 
pendent  bodies,  which,  however,  remain  paired’ m  fhc’ telophase,  and 
occupy  a  great  variety  of  positions  in  regard,  ,to  -^ach  other.  The 
homotypic  chromosomes  thus  assume  rnany  ot  the.,  characteristic 
shapes  which  are  usually  observed  in  the  heterpvy pic  chromosomes  of 
other  forms,  such  as  X,  Y,  V,  H,  etc.  The  failure  of  the  heterotypic 
bivalents  to  form  these  shapes  is  due  partly  to  the  weaker  attraction 
between  the  members  of  a  pair,  but  largely  to  a  difference  in  their 
shape,  each  member  of  a  pair  being  usually  more  rounded  in  the 
heterotypic  and  more  elongated  and  rodlike  during  the  stages  between 
the  two  mitoses. 

The  telophase  of  the  heterotypic  mitosis  is  one  of  the  best  stages  for 
counting  the  chromosomes,  as  they  are  distributed  at  equal  intervals 
around  the  periphery  of  the  nucleus,  no  two  ever  being  in  contact  and 
the  halves  of  each  (bivalent)  chromosome  rarely  separating.  The 
chromosomes  now  evidently  repel  each  other,  while  the  halves  of 
each  chromosome  attract  each  other  rather  strongly.  The  halves  of 
these  bivalent  chromosomes  are  usually  short  rods,  but  they  may  be 
dumb-bell  or  hour-glass  shaped,  or  nearly  globular,  as  previously 
mentioned  (12).  Sometimes,  however,  this  split  fails  completely  to 
occur  in  the  anaphase,  the  daughter  chromosomes  remaining  single 
and  globular  or  somewhat  elongated  {fig.  jp).  These  telophase 
stages  and  the  prophases  of  the  homotypic  mitosis  will  be  taken  up 
in  detail  in  a  paper  dealing  with  different  forms.  These  results, 
therefore,  will  not  be  duplicated  here,  but  a  brief  statement  of  the 
events  of  the  second  mitosis  will  be  given. 


HOMOTYPIC  MITOSIS 

In  the  telophase  of  the  heterotypic  mitosis  the  nuclei  never  pass 
into  the  resting  condition  and  the  chromosomes  never  lose  their 
identity  completely,  though  they  spread  out  and  anastomose  with 
each  other  more  or  less.  Nucleoli  are  formed,  as  previously  described 
(ii).  These  stages  between  the  two  mitoses  last  for  some  time,  but 
the  events  of  the  second  mitosis  are  passed  through  very  quickly.  The 
two  homotypic  spindles  are  formed  simultaneously  and  their  axes  are 


i8 


BOTANICAL  GAZETTE 


[JULY 


at  various  angles  to  each  other.  Spindle  formation  is  the  same  as  for 
the  heterotypic  mitosis,  except  that  the  spindles,  are  smaller.  In 
regard  to  the  chi^omatln,’ Suffice  it  at  present  to  say  that  the  chromo¬ 
somes -of  ^he  hOmotypic '  prophase  show  the  same  general  types  and 
are  often  identical  in'  appearance  with  those  of  the  heterotypic  telo¬ 
phase.  '  ■  ''Ki'e’recan.  be  n'o'dOubt  that  the  bivalent  bodies  which  appear 
on  the  homotypic.  spindle"  are  the  same  bodies  that  were  present  in  the 
telophase  of  the  heterotypic.  Fig.  41  shows  an  early  anaphase  of  the 
second  mitosis,  the  members  of  each  pair  having  just  separated.  One 
of  the  small  nucleoli  appears  by  one  of  the  spindles. 

IRREGULARITIES 

In  fig.  jg  spindle  fibers  are  seen  in  the  cytoplasm  by  the  side  of  the 
spindle  in  anaphase.  This  may  be  connected  with  a  condition  which 
is  illustrated  in  fig.  40.  Six  such  cases  were  observed  in  which  a 
regular  spindle  occurred  at  the  side  of  the  mother  cell  instead  of 
between  the  daughter  nuclei,  after  the  partial  or  complete  disappear¬ 
ance  of  the  heterotypic  spindle.  Some  of  these  cases  were  in  the  telo¬ 
phase  of  the  heterotypic  spindle  {fig.  40) ;  others  were  in  the  prophase 
of  the  homotypic.  In  these  cases  the  spindles  were  regularly  formed 
and  rather  sharp-pointed  and  occupied  the  same  position  at  the  side 
of  the  cell;  of  course  they  contained  no  chromosomes.  The  method 
of  their  origin  is  unknown,  but  it  seems  probable  that  they  are  con¬ 
nected  with  the  condition  observed  in  fig.  jg.  Mother  cells  which 
probably  indicate  an  intermediate  condition,  in  which  irregularly 
arranged  fibers  were  found  at  the  side  of  the  cell,  were  occasionally 
observed.  They  may  merely  indicate  a  persistence  of  the  kinoplasm 
of  the  heterotypic  spindle  after  its  function  has  ceased,  but  their 
structure  appeared  remarkably  definite  in  most  of  the  cases  observed. 

A  single  case  of  extra  nuclei  in  the  pollen  tetrad  was  observed  in 
O.  rubrinervis.  These  have  been  previously  described  in  O.  lata 
(ii),  where  they  are  common  occurrences  in  connection  with  pollen 
degeneration.  The  single  case  observed  in  O.  rubrinervis  is  sketched 
in  fig.  42.  '  Two  small  nuclei  are  present  in  addition  to  the  four 
larger  ones  composing  the  tetrad.  The  nuclei  had  passed  too  far 
into  the  resting  condition  to  count  the  chromosomes  in  each 
nucleus. 


1908] 


CjATES— REDUCTION  IN  OENOTHERA 


19 


POLLEN  DEGENERATION 

The  general  question  of  pollen  degeneration  in  Oenothera  is  an 
interesting  one.  It  reaches  its  extreme  expression  in  O.  lata,  which 
is  usually  completely  sterile  in  this  regard,  and  in  which  I  have 
already  shown  (ii)  that  irregularities  occur  during  the  reduction 
divisions  similar  to  those  found  in  sterile  hybrids.  The  question  of 
sterility  is  evidently,  as  Tischler  (32)  suggests,  a  relative  one. 

In  O.  rubrinervis  one  is  led  from  a  gross  examination  to  judge 
that  the  pollen  production  is  copious  and  probably  equal  to  that  of 
O.  Lamarckiana  itself,  but  in  reality  many  of  the  pollen  mother  cells 
fail  to  complete  their  divisions.  From  an  examination  of  sections  of 
anthers  of  O.  rubrinervis  it  is  found  that  in  some  loculi  a  large  number 
or  perhaps  nearly  all  the  mother  cells  may  be  degenerating  in  the 
synapsis  stage.  Frequently  the  cells  are  flattened  and  distorted, 
appearing  pressed  together  for  lack  of  space  in  the  loculus.  The 
chromatic  contents  of  such  cells  often  form  a  dense  irregular  mass,  or 
their  nuclei  may  be  in  normal  synapsis  or  mitosis,  notwithstanding  the 
distorted  shape  of  the  cell;  while  still  other  cells  of  the  same  loculus 
may  be  entirely  normal.  Even  earlier,  in  the  archesporial  stage,  the 
tapetal  cells  in  many  sections  were  found  to  be  breaking  down,  as  in 
O.  lata  (ii).  No  indications  of  degeneration  have  yet  been  observed 
in  mother  cells  of  O.  Lamarckiana,  and  very  few  in  the  tapetum. 

The  percentage  of  mother  cells  which  thus  degenerate  in  O. 
rubrinervis  was  not  determined.  Tischler  (32)  suggests  that  the 
causes  of  sterility  in  mutants  are  the  same  as  those  in  hybrids  and  in 
plants  under  cultivation.  This  general  cause  he  designates  as  a 
disturbance  or  derangement  of  the  constitution  of  the  idioplasm,  which 
he  thinks  has  taken  place  in  the  production  of  mutants  as  well  as  in 
hybrids  and  under  the  conditions  of  cultivation. 

PROTOPLASMIC  CONNECTIONS 

It  is  an  interesting  fact  that  large  and  rather  conspicuous  proto¬ 
plasmic  connections  are  found  between  the  pollen  mother  cells  in 
O.  rubrinervis.  They  are  usually  quite  easily  seen  and  it  is  probable 
that  they  are  always  present.  They  consist  of  delicate  strings  or 
threads  of  cytoplasm  connecting  adjacent  mother  cells.  In  size  they 
vary  greatly,  from  the  delicacy  of  a  spindle  fiber  to  a  coarse  thread  or 


20 


BOTANICAL  GAZETTE 


[JULY 


strand  connecting  the  cells  {figs.  45,  46).  When  the  cytoplasm  has 
shrunken  slightly  away  from  the  cell  wall  they  are  particularly  clearly 
observable.  These  connections  appear  to  be  in  all  cases  between 
mother  cells,  and  in  no  case  have  they  been  observed  between  the 
mother  cells  and  the  tapetum.  Generally  one  such  strand  is  seen 
connecting  two  cells,  but  not  infrequently  there  are  two  or  three  or 
occasionally  even  more.  There  is  no  constriction  or  change  in  the 
nature  of  the  connective  as  it  passes  through  the  cell  wall.  These 
connections  are  even  larger  and  more  conspicuous  in  O.  gigas,  where 
the  mother  cells  are  also  much  larger.  They  have  not  been  observed 
in  O.  Lamarckiana  or  the  other  forms,  but  they  doubtless  occur  in  all, 
being  probably  smaller  and  more  inconspicuous  in  some. 

Discussion 

The  method  of  reduction  described  in  this  paper  at  once  raises  a 
number  of  questions  of  prime  importance  from  the  cytological  stand¬ 
point,  as  well  as  from  that  of  the  relation  subsisting  between  heredi¬ 
tary  and  cytological  phenomena.  A  discussion  of  all  these  features 
will  not  be  attempted  at  this  time,  the  intention  of  the  writer  being 
merely  to  indicate  the  general  directions  in  which  the  facts  point  and 
the  possible  bearing  which  these  data  may  be  found  to  have  on  the 
problems  connected  with  the  phenomena  of  mutation  in  Oenothera. 
A  fuller  discussion  of  these  subjects  is  reserved  for  a  future  time,  after 
the  presentation  of  further  data.  In  the  present  communication 
reference  will  be  made  only  to  the  most  recent  papers  on  reduction  in 
plants,  the  purpose  not  being  a  review  of  the  literature,  or  a  dis¬ 
cussion  of  present  views,  except  in  so  far  as  they  bear  directly  on  the 
matter  in  hand. 

The  recent  accounts  of  reduction  in  plants,  given  by  Berghs 
(3>  4>  5»  6),  Gregoire  (16),  Strasburger  (31),  Allen  (i,  2), 
Miyake  (18),  Overton  (22),  Rosenberg  (25),  Yamanouchi  (33), 
and  others,  have  agreed  in  so  far  as  the  following  general  course  of 
events  is  concerned :  In  synapsis  a  pairing  of  homologous  maternal 
and  paternal  elements  occurs  either  in  the  form  of  gamosomes  (Stras¬ 
burger  and  Miyake),  prochromosomes  (Overton),  or  parallel 
threads  (Allen,  Rosenberg,  Gregoire,  Berghs,  Cardiff  7,  and 
Yamanouchi).  In  every  case  two  parallel  threads  result,  which  unite 


1908] 


GATES— REDUCTION  IN  OENOTHERA 


21 


more  or  less  intimately  about  the  time  of  synapsis  or  later.  After  the 
events  of  synapsis,  a  longitudinal  split  reappears  in  the  thickened 
spirem  threads,  this  split  representing  the  line  of  approximation  of  the 
two  original  spirems.  Transverse  segmentation  into  pairs  of  chro¬ 
mosomes,  which  are  believed  to  be  homologous  somatic  chromosomes 
of  maternal  and  paternal  origin,  then  takes  place.  The  halves  of  these 
bivalent  chromosomes,  which  lie  side  by  side,  are  then  distributed 
in  the  heterotypic  mitosis,  which  is  thus  a  reduction  division.  In 
the  anaphase  of  the  heterotypic  mitosis  a  longitudinal  split  appears  in 
the  daughter  chromosomes,  which  is  regarded  as  a  premature  split 
for  the  homotypic  mitosis,  the  latter  being  thus  an  equation  division. 
The  persistency  with  which  this  general  account  has  been  given,  not¬ 
withstanding  differences  in  detail,  particularly  preceding  and  during 
synapsis,  leads  the  writer  to  the  belief  that  it  is  probably  correct  in  its 
main  outlines,  at  least  in  many  of  the  forms  described.  This  being 
judged  to  be  the  case,  every  effort  was  made  to  bring  the  account  in 
Oenothera  into  harmony  with  this  general  course  of  events  but  with¬ 
out  success,  for  Oenothera  is  found  to  deviate  in  some  important 
particulars,  as  is  already  evident  from  the  description. 

Another  general  account  of  reduction  in  plants,  which  was  adhered 
to  by  Strasburger  as  late  as  1904  (30),  and  has  been  held  notably 
by  Farmer  and  Moore  (8,  9),  Farmer  and  Shove  (10),  Schaffner 
(28),  Mottier  (19,  20),  and  others,  to  mention  only  a  few  of  the  recent 
papers,  is  in  general  as  follows :  The  split  in  the  spirem  which  occurs 
at  about  the  time  of  synapsis  is  a  true  split,  such  as  may  occur  in  the 
prophase  of  somatic  mitoses,  and  is  not  preceded  by  a  pairing  of 
parallel  threads,  but  the  thread  is  single  from  the  beginning.  This 
split  afterward  closes  up  as  the  thread  shortens  and  thickens  after 
synapsis,  and  the  single  spirem  so  formed  segments  usually  into  the 
reduced  number  of  chromosomes,  which  are  thus  arranged  successively 
end  to  end.  Each  such  bivalent  chromosome  thus  consists  of  two 
halves  arranged  end  to  end,  not  side  by  side,  and  the  heterotypic 
mitosis  thus  separates  successive  whole  chromosomes  on  the  spirem, 
being  therefore,  as  in  the  other  account,  a  reduction  division.  The 
split  which  appears  in  the  anaphase  of  this  mitosis  is  interpreted  as  a 
reappearance  of  the  earlier  longitudinal  split  of  the  spirem.  The 
homotypic  mitosis  is  therefore  an  equation  or  longitudinal  division. 


t 


22 


BOTANICAL  GAZETTE 


[JULY 


There  are  of  course  minor  differences  in  these  accounts,  Schaffner 
(28)  stating,  for  example,  that  in  Lilium  tigrinum  there  is  a  splitting 
of  granules  in  the  spirem,  but  the  linin  thread  remains  single.  Differ¬ 
ences  of  opinion  are  also  expressed  regarding  the  arrangement  of  the 
loops  of  the  spirem  before  segmentation,  and  their  relation  to  the 
chromosomes  formed. 

These  two  general  schemes  agree  that  the  heterotypic  mitosis  is 
a  reduction  division  separating  whole  somatic  chromosomes,  while 
the  second  division  is  longitudinal.  The  essence  of  the  distinction  is 
that  the  first  view  regards  the  chromosome  bivalents  as  formed  by  a 
side-by-side  union  of  homologous  chromosomes  through  the  medium 
of  parallel  threads,  while  the  second  view  holds  to  an  end-to-end 
union.  It  will  be  seen  that,  omitting  the  points  which  are  left  undeter¬ 
mined,  the  account  in  Oenothera  corresponds  more  nearly  with  the 
latter  scheme  than  with  the  former,  though  differing  in  some  respects 
from  both.  Rosenberg  (25),  from  a  comparison  of  forms  having 
long  and  short  chromosomes,  has  attempted  to  harmonize  the  latter 
view  with  the  former.  He  examined  Listera,  Tanacetum,  Drosera, 
and  Arum,  and  found  that,  for  example  in  Drosera,  which  has  short 
definitive  chromosomes  much  like  those  of  Oenothera,  the  spirem 
first  segmented  into  long  twisted  chromosomes  lying  in  pairs  with 
their  long  axes  parallel.  Later,  as  they  condensed  into  the  short, 
rounded  definitive  chromosomes,  they  frequently  swung  around  end 
to  end,  so  that  an  observer  seeing  only  the  later  stage  would  conclude 
that  they  had  been  arranged  tandem  on  the  spirem  at  the  time  of  their 
origin.  Similar  conditions  were  sometimes  observed  in  Listera. 
I  think  my  figs.  22-28  make  it  evident  that  this  explanation  will  not 
apply  to  Oenothera.  The  chromosomes  in  Oenothera  do  not  undergo 
any  such  great  amount  of  condensation,  but  are  already  thick,  heavy 
bodies  when  first  formed  from  segmentation  of  the  spirem  {fig.  24). 
Their  diameter  at  this  time  is  about  the  same  as  that  of  the  spirem 
just  previous  to  segmentation,  as  is  shown  by  comparing  figs.  22  and 

with  figs.  24  and  26.  The  fact  that  as  many  as  eight  or  more 
chromosomes  may  be  found  forming  a  single  connected  chain  {fig.  26) 
also  renders  this  explanation  impossible. 

Miyake  (18)  finds  that  after  the  pairing  of  elements  in  synapsis 
(the  exact  method  of  this  pairing  need  not  be  entered  into  here)  in 


1908] 


GATES— REDUCTION  IN  OENOTHERA 


23 


Galtonia  and  Tradescantia,  a  longitudinal  split  appears  in  the  thick¬ 
ened  thread,  and  the  double  spirem  thus  formed  breaks  transversely 
into  the  reduced  number  of  chromosome  pairs.  Later,  in  these 
forms,  a  secondary  union  between  the  chromosomes  is  claimed  to  take 
place,  forming  a  single  connected  chain  of  chromosomes  (as  in  Oeno¬ 
thera).  Sometimes  a  pair  of  chromosomes  lies  free  by  itself  at  this 
time.  Then  by  further  shortening  the  chromosomes  of  Galtonia 
again  fall  apart  into  pairs,  though  in  Tradescantia  they  frequently 
remain  connected  even  after  spindle  formation.  The  apparent 
similarity  of  the  chromosome  chain  thus  described  by  Miyake  in 
Galtonia  to  the  condition  in  Oenothera,  led  the  writer  to  make  an 
endeavor  to  harmonize  the  two  accounts.  But  instead  of  this,  all 
the  evidence  obtained  from  a  critical  study  of  the  stages  concerned 
shows  that  in  Oenothera  a  single  very  thick  spirem  breaks  transversely 
into  the  sporophyte  number  of  chromosomes.  A  critical  examination 
of  jigs.  22-28  will  make  it  clear,  I  think,  that  we  are  following  the 
progressive  segmentation  of  a  single  spirem,  and  there  is  no  room 
for  stages  between,  in  which  a  double  spirem  breaks  into  two  parallel 
series  of  chromosomes.  Moreover,  it  is  hardly  likely  that  secondary 
fusions  between  chromosomes  would  take  place  to  such  an  extent  as  is 
shown  in  jigs.  and  24.  In  nuclei  such  as  'jig.  20,  in  which  a  pair  of 
chromosomes  is  cut  off  prematurely  from  the  spirem  while  still  in  the 
second  contraction,  they  are  invariably  connected  at  one  end  and 
rarely,  if  ever,  at  the  other  (though  sometimes  the  close  approximation 
of  the  latter  ends  may  give  the  false  appearance  of  a  ring).  This 
would  not  be  the  case  if  they  came  from  separate  paired  threads 
merely  lying  side  by  side,  so  that  this  connection  shows  them  to  have 
been  really  successive  on  the  spirem.  From  this  evidence  the  writer 
cannot  see  how  anything  except  a  distortion  of  the  facts  can  lead  to  the 
assumption  in  Oenothera  of  two  parallel  threads  breaking  into  chro¬ 
mosomes.  Hence  the  conclusion  is  that  the  double  threads  appear¬ 
ing  in  the  stage  represented  by  jig.  17  have  united  to  form  a  single 
thread,  which  then  breaks  transversely  into  the  sporophyte  number 
of  chromosomes. 

This  corresponds  fairly  well  with  Strasburger’s  1904  (30)  account 
of  the  post-synaptic  stages  in  Galtonia,  and  suggests  to  the  writer 
that  perhaps  after  all  the  earlier  account  may  be  nearer  the  facts. 


24 


BOTANICAL  GAZETTE 


[JULY 


SO  far  as  the  points  here  under  discussion  are  concerned,  than  the 
paper  of  1905  (31).  The  close  similarity  of  the  conditions  in  Galtonia 
and  Tradescantia  during  diakinesis  to  those  in  Oenothera  suggests 
that  they  may  be  found  finally  to  conform  to  Oenothera  in  these  later 
stages.  Whether  or  not  this  will  be  found  to  be  the  case,  we  must 
conclude  that  in  Oenothera  the  longitudinal  fission  in  the  spirem 
(however  it  originated)  closes  up,  and  that  after  the  second  contrac¬ 
tion,  or  during  it,  the  thick  thread  segments  into  the  sporophyte 
number  of  chromosomes.  Since  this  diverges  in  important  respects 
from  nearly  all  the  recent  accounts  of  reduction  in  plants,  the  con¬ 
clusion  is  that  reduction  probably  takes  place  differently  in  different 
plants.  Whether  or  not  the  results  are  different  from  the  standpoint 
of  a  qualitative  distribution  will  not  be  discussed  now.  The  writer 
believes  the  above  conclusions  to  be  necessary,  despite  the  fact  that 
authors  have  reached  different  conclusions  in  regard  to  the  same 
plant,  particularly  in  such  cases  as  Lilium  and  Podophyllum. 

The  next  important  point  which  requires  discussion  and  which 
was  left  undecided  in  the  statement  of  observations,  is  in  regard  to 
whether  the  double  thread  observed  after  synapsis  arises  from  an 
approximation  of  parallel  filaments  or  through  a  primary  split  in  the 
thread.  It  may  be  well  to  examine  the  results  which  follow  from 
either  assumption.  The  writer  hopes  later  to  determine  more  defi¬ 
nitely  this  difficult  matter.  On  the  first  assumption  of  a  lateral 
approximation  in  synapsis  of  two  spirems  representing  respectively 
the  maternal  and  paternal  chromosomes,  we  should  expect  the  double 
thread  so  formed  to  segment  into  the  reduced  number  of  chromosome 
pairs,  in  order  to  conform  to  the  current  account  in  forms  in  which 
there  is  a  pairing  of  spirems,  for  example  Allen  (i),  Gregoire  (16), 
and  Yamanouchi  (33).  Instead,  however,  the  spirem  segments 
into  the  unreduced  number  of  bodies.  We  may  still  assume  that 
each  of  these  bodies  consists  of  maternal  and  paternal  longitudinal 
halves  still  closely  held  together  and  resulting  from  a  previous  approxi¬ 
mation.  According  to  this  view  the  first  mitosis  would  separate 
bodies  which  were  arranged  successively  on  the  spirem,  while  the 
second  mitosis  would  separate  the  maternal  and  paternal  halves  of 
these  bodies.  The  reason  for  such  a  result  would  be  that  the  maternal 
and  paternal  spirems  remained  closely  fused  after  pairing,  so  that 


1908] 


GATES— REDUCTION  IN  OF.NOTHERA 


25 


their  elements  were,  separated  in  the  second  mitosis  instead  of  the 
first.  This  view  is  scarcely  admissible  for  several  reasons.  In  the 
first  place,  on  this  hypothesis  transverse  segmentation  of  the  spirem 
must  have  taken  place  not  only  between  the  (bivalent)  chromosomes 
but  also  in  the  middle  of  each  chromosome,  in  order  to  give  a  chain 
of  fourteen  bodies.  Such  a  segmentation  seems  unlikely.  Another 
possible  explanation  would  be  that  the  chromosomes  have  lost  their 
identity  during  synapsis,  and  that  the  bodies  we  are  dealing  with  now 
are  new  arrangements  of  the  chromatic  material,  irrespective  of  the 
somatic  chromosomes.  Many  considerations,  however,  strongly  sup¬ 
port  the  belief  that  these  bodies  really  represent  the  somatic  chro¬ 
mosomes.  The  facts  so  far  educed  in  Oenothera,  in  the  opinion  of 
the  writer,  all  favor  the  hypothesis  of  the  separate  existence  and 
genetic  continuity  of  the  chromosomes  from  one  generation  to  another. 
In  this  connection  may  be  cited  certain  plants  from  the  Fi  of  O.  lata 
XO.  gigcLS,  which  as  stated  elsewhere  (14)  have  21  chromosomes  as 
somatic  number,  10  of  which  regularly  go  to  one  pole  of  the  hetero¬ 
typic  spindle  and  ii  to  the  other.  Occasionally,  however,  the  segre¬ 
gated  numbers  of  chromosomes  are  12  and  9,  one  chromosome  having 
gone  to  the  wrong  pole  of  the  spindle.  In  this  hybrid  7  of  the  chro¬ 
mosomes  are  maternal  and  14  paternal.  If  in  this  case  there  were  a 
pairing  of  maternal  and  paternal  spirems,  it  is  difficult  to  see  how  it 
could  be  accomplished  and  result  in  the  distribution  of  chromosomes 
in  the  heterotypic  mitosis  already  stated. 

It  will  be  instructive  to  compare  the  chromosome  history  in  this 
cross  with  the  often-quoted  condition  found  by  Rosenberg  (23,  24) 
in  Drosera  longijoliaXD.  rotundi folia.  D.  rotundijolia  has  10  chro¬ 
mosomes  and  D.  longifolia  20,  as  the  gametophyte  number.  The 
hybrid  naturally  has  30  chromosomes  in  its  sporophyte  tissues, 
but  in  diakinesis  20  chromosome  bodies  appear,  10  of  which  are 
double,  consisting  of  a  larger  and  a  smaller  half,  while  the  remaining 
10  are  the  unpaired  (smaller)  longifolia  chromosomes.  The  larger 
and  smaller  halves  of  the  10  bivalents  separate  and  pass  regularly 
to  the  poles  of  the  heterotypic  spindle,  but  the  unpaired  chromosomes 
are  irregularly  distributed  or  left  out  of  the  daughter  nuclei.  Later 
the  pollen  deteriorates.  This  result  is  strikingly  different  from  that 
in  the  Oenothera  hybrid,  and,  while  perfectly  in  harmony  with  the 


26 


BOTANICAL  GAZETTE 


[JULY 


idea  of  the  pairing  of  threads  in  synapsis  in  Drosera,  makes  it  highly 
probable,  and  in  fact  necessary,  that  the  method  of  reduction  in  the 
Oenothera  hybrid  be  different.  This  is  a  strong  argument  not 
only  against  pairing  of  maternal  and  paternal  spirems  in  Oenothera, 
but  in  favor  of  the  probability  that  reduction  takes  place  in  diverse 
ways  in  the  two  genera.  A  considerable  amount  of  time  has  already 
been  devoted  to  the  study  of  reduction  in  this  Oenothera  hybrid,  and 
an  account  will  be  published  later.  So  far  as  observed  it  shows  no 
differences  in  method  from  the  account  given  here  for  the  pure  races. 

The  hypothesis  of  the  pairing  of  parental  spirems  in  synapsis  in 
Oenothera  being  thus  rejected,  the  other  alternative  remains,  namely, 
that  the  double  spirem  results  from  a  split ;  and  this  appears  to  satisfy 
all  the  facts.  The  observations  have  already  shown  that  the  spirem 
segments  into  a  single  chain  of  chromosomes.  The  description  of 
events  in  Oenothera  from  synapsis  on  thus  agrees  in  outline  with  the 
1904  account  of  Strasburger  (30)  in  Galtonia,  and  in  general  also 
with  that  of  Farmer  and  Moore  (9)  in  Lilium,  Osmunda,  Psilotum, 
and  Aneura,  Farmer  and  Shove  (10)  in  Tradescantia,  and  Mottier 
(19,  20)  in  Lilium,  Podophyllum,  and  Tradescantia.  The  belief  of 
the  writer  is  that  some  of  these  forms  will  be  found  to  correspond 
more  nearly  with  the  account  which  involves  a  pairing  of  threads,  and 
some  with  the  account  involving  only  a  split. 

Another  important  matter  which  requires  mention  at  this  time 
is  the  nature  of  the  chromosome  distribution  which  takes  place  on  the 
heterotypic  spindle  in  Oenothera.  As  already  observed,  the  chromo¬ 
somes  even  during  spindle  formation  are  frequently  unpaired.  This 
appears  to  be  due  to  the  weakness  of  the  mutual  attraction  which 
ordinarily  leads  to  pairing.  Granting  that  homologous  maternal  and 
paternal  chromosomes  unite  when  pairing  takes  place,  what  are  the 
possibilities  regarding  the  unpaired  chromosomes  ?  Pairing  insures 
ordinarily  that  the  members  of  the  pair  will  proceed  to  opposite 
poles  of  the  spindle,  and  hence  that  the  homologous  maternal  and 
paternal  elements  will  enter  different  nuclei.  There  is  no  such 
certainty  in  the  distribution  of  the  unpaired  chromosomes,  so  that  it 
might  be  expected  that  in  certain  cases  both  members  of  a  pair  would 
enter  the  same  daughter  nucleus.  It  is  important  to  note  that  this 
result  is  entirely  independent  of  the  origin  of  these  chromosome 


1908] 


CiATES— REDUCTION  IN  OENOTHERA 


27 


pairs,  whether  from  an  end-to-end  or  side-by-side  union  of  somatic 
chromosomes,  or  in  any  other  manner,  so  that  this  question  holds  no 
necessary  relation  to  the  method  of  reduction.  On  the  common 
cytological  assumption  that  the  chromosomes  are  qualitatively 
different  (which  has  apparently  been  shown  to  be  a  fact  in  certain 
well-known  cases  in  animals,  that  need  not*  be  cited),  germ  cells 
would  occasionally  arise  lacking  both  members  of  a  pair^  and  hence 
lacking  the  possibility  of  developing  certain  qualities.  In  this  manner 
it  is  conceivable  that  a  series  of  types  might  arise  from  the  parent 
O.  Laniarckiana,  each  lacking  the  possibility  of  developing  a  certain 
group  of  characters  possessed  by  O.  Lamarckiana. 

On  this  view,  which  is  suggested  merely  as  a  tentative  hypothesis, 
we  would  have  in  the  mutations  of  O.  Lamarckiana  an  analytical 
process  in  which  a  series  of  types  arises  from  the  parent  form,  each 
lacking  in  a  different  group  of  qualities  or  capacities  which  the  parent 
form  possessed.  This  does  not  apply  to  O.  gigas,  however,  which  will 
be  taken  up  at  another  time.  The  further  bearings  of  this  hypothesis 
on  the  mutation  theory  of  DeVries  will  not  be  followed  up  in  this 
discussion,  but  it  may  be  pointed  out  here  that  such  a  hypothesis 
accounts  for  the  absence  of  reversions  of  the  mutants  to  O.  Lamarcki¬ 
ana,  and  it  may  also  account  for  some  of  the  peculiarities  of 
hybridization  among  the  Oenothera  mutants.  I  should  therefore  sug¬ 
gest  that  there  may  be  a  relation  between  the  type  of  reduction  in  any 
organism  and  its  variation  and  hybridization  phenomena. 

In  Galtonia  and  probably  also  in  Tradescantia  there  are  apparently 
the  same  possibilities  that  both  chromosomes  of  a  pair  may  occasion¬ 
ally  enter  the  same  daughter  nucleus.  In  other  plant  forms  studied 
the  attraction  between  chromosomes  seems  to  be  strong  enough  to 
keep  the  members  of  a  pair  together  until  their  separation  in  the 
anaphase  of  the  heterotypic  mitosis.  The  segregation  of  the  members 
of  a  pair  into  separate  germ  cells  is  thus  insured.  In  cases  where, 
as  in  Oenothera,  the  members  of  a  pair  do  not  always  remain  in 
contact,  but  are  loosely  arranged  on  the  spindle,  such  a  result  as 
already  suggested  seems  certain  to  occur  in  certain  instances. 

It  has  already  been  mentioned  that  occasionally  one  chromosome 
goes  to  the  wrong  pole  of  the  heterotypic  spindle.  This  is  found  to 
be  the  case  particularly  in  the  hybrids,  for  example,  in  the  O.  Lamarcki- 


t 


28 


BOTANICAL  GAZETTE 


[JULY 


ana  plants  from  the  Fi  of  O.  lataXO.  Lamarckiana  (13),  in  which 
sometimes  eight  chromosomes  pass  to  one  pole  and  six  to  the  other; 
but  it  may  also  occur  rarely  in  the  pure  races.  This  matter  was 
briefly  discussed  elsewhere  (14).  Assuming  that  the  14  chromosomes 
are  in  two  similar  sets  of  7  each,  and  that  homologous  members  of 
these  sets  conjugate  except  when  there  is  a  failure  to  pair,  then  when 
8  chromosomes  go  to  one  pole  and  6  to  the  other,  both  members  of  one 
of  the  pairs  must  have  gone  to  the  same  pole.  This  probably  takes 
place  in  cases  where  such  members  were  unconjugated,  for  the 
purpose,  or  at  any  rate,  the  result  of  the  pairing  is  in  ordinary  cases 
that  one  member  of  every  pair  shall  be  distributed  to  each  pole.  If, 
while  two  members  of  one  pair  thus  go  to  one  pole,  the  second  member 
of  another  pair  goes  to  the  other  pole,  we  should  have  an  equal 
numerical  distribution  of  chromosomes,  but  one  daughter  group  would 
be  lacking  both  members  of  one  pair  and  the  other  would  be  lacking 
both  members  of  another  pair.  It  is  highly  probable  that  such  a 
distribution  occasionally  takes  place,  though  it  would  be  less  common 
than  the  case,  already  proved,  where  the  members  of  a  single  pair  are 
unilaterally  distributed.  It  should  be  borne  in  mind  that  such  cases 
are  most  likely  to  occur,  not  when  the  members  of  a  pair  are  con¬ 
jugated,  but  when  they  lie  separately  in  diakinesis  and  on  the  spindle. 

Miss  Lutz  (17),  from  an  examination  of  root  tips,  states  that  she 
has  observed  several  individuals  belonging  to  different  strains  having 
15  chromosomes  instead  of  14.  This  is  to  be  anticipated  from  the 
irregularities  in  chromosome  distribution  in  reduction  already  men¬ 
tioned.  I  have  observed  one  such  case  in  O.  lataXO.  gigas  (14)  — 
a  certain  plant  having  20  chromosomes  instead  of  21.  All  the  plants 
of  O.  lata  (12)  and  O.  nanella  (13)  thus  far  examined  by  me  had  14 
chromosomes,  while  Miss  Lutz  (17)  finds  in  root  tips  some  O.  lata 
plants  with  14  and  also  some  with  15  or  she  thinks  possibly  16  chro¬ 
mosomes.  She  reports  finding  two  O.  fianella  plants  with  14  chro¬ 
mosomes  and  one  with  15.  Two  O.  albida  seedlings  are  said  to  have 
15  chromosomes  and  two  O.  oblonga  plants  15,  while  a  third  has  14. 
Disregarding  the  possibility  that  these  results  might  be  due  to  the 
well-known  variation  in  chromosome  numbers  in  root  tips,  they  are 
such  as  would  be  likely  to  arise  in  different  individuals  from  the 
cytological  irregularities  I  have  already  described.  Whether  there 


GATES— REDUCTION  IN  OENOTHERA 


29 


1908] 

are  external  differences  between  the  plants  having  14  chromosomes 
and  those  of  the  same  race  having  15,  is  as  yet  unknown.  But  it  is 
quite  conceivable  that  no  such  differences  will  be  found,  for  if  the 
sporophyte  chromosomes  consist  of  two  complete  sets  (and  for  a 
variety  of  reasons  this  seems  the  only  tenable  view  at  the  present  time 
if  we  assume  qualitative  differences  at  all),  the  presence  of  an  addi¬ 
tional  chromosome,  which  is  already  present  in  duplicate,  would 
scarcely  be  expected  visibly  to  affect  the  plant. 

Rosenberg  (26)  has  found  an  analogous  situation  in  Hieracium. 
For  example,  H.  excellens'KH .  Pilosella  gives  hybrids  with  different 
numbers  of  chromosomes.  This  he  ascribes  to  the  fact  that  the  eggs 
of  H.  excellens  differ  in  their  numbers  of  chromosomes,  which  he  finds 
is  due  to  irregularities  in  chromosome  distribution  during  the  reduction 
divisions.  The  writer  has  pointed  out  elsewhere  (12)  certain  similari¬ 
ties  between  the  hybridization  phenomena  in  Hieracium  and  Oeno¬ 
thera,  and  this  seems  to  be  a  further  similarity  between  the  two  genera.' 

Rosenberg  (27)  has  since  shown  that  H.  excellens  produces  three 
kinds  of  embryo  sacs:  (i)  Normal  embryo  sacs  which  require  fertili¬ 
zation  for  their  development.  These  are  presumably  the  only  ones 
which  can  be  hybridized.  The  egg  cells  in  these  sacs  vary  in  their 
number  of  chromosomes  owing  to  the  fact  that  some  of  the  chromo¬ 
somes,  lacking  in  “affinity,”  remain  univalent  (that  is,  fail  to  pair) 
during  the  heterotypic  mitosis  and  are  irregularly  distributed.  It  is 
evident  that  this  lack  of  affinity  between  chromosomes  is  similar  to  that 
in  Oenothera.  (2)  In  rare  cases  apogamous  embryo  sacs  are  formed 
after  a  single  division  of  the  megaspore  mother  cell,  and  without 
reduction.  (3)  More  frequently  the  condition  occurs  which  Rosen¬ 
berg  calls  apospory,  in  which  tetrad  formation  takes  place  and  then 
an  adjacent  cell  of  the  nucellus  enlarges,  displaces  the  tetrad,  and 
forms  an  embryo  sac  without  reduction. 

Summary 

In  conclusion  a  brief  summary  of  the  facts  and  considerations 
here  presented  will  be  useful. 

I.  In  Oenothera  the  heterotypic  mitosis  is  a  reduction  division, 
separating  whole  chromosomes  which  lie  successively  on  the  spirem. 
The  homotypic  mitosis  is  an  equation  division,  separating  the  longi- 


30 


BOTANICAL  GAZETTE 


[JULY 


tudinal  halves  of  the  daughter  chromosomes  of  the  heterotypic 
mitosis.  Whether  an  approximation  of  threads  or  a  split  in  a  single 
thread  occurs  in  synapsis  was  not  determined  with  certainty  from  the 
observations,  but  various  considerations  lead  to  the  belief  that  in 
Oenothera  the  doubling  is  due  to  a  split  which  closes  up  later,  rather 
than  to  an  approximation  of  separate  spirems. 

2.  The  conclusion  that  the  method  of  reduction  probably  differs 
in  different  genera  is  based  on  two  considerations:  (i)  the  fact  that 
in  most  of  the  recent  accounts  of  synapsis  and  reduction  in  plants 
a  side-by-side  pairing  of  chromosomes  from  maternal  and  paternal 
spirems  is  described,  while  in  Oenothera  the  members  of  a  pair  are 
arranged  end  to  end  on  a  single  spirem;  and  (2)  on  differences  in 
chromosome  distribution  during  reduction  in  certain  hybrids  of 
Drosera  and  of  Oenothera  (see  p.  25).  If  reduction  took  place  in 
the  same  manner  in  both  genera,  the  chromosome  distribution  during 
reduction  in  these  hybrids  with  reference  to  the  parental  chromosome 
numbers  should  be  the  same  in  both,  but  this  is  not  the  case. 

3.  Pairing  between  the  definitive  chromosomes  during  diakinesis 
and  the  prophase  of  the  heterotypic  mitosis  does  not  always  take  place, 
owing  to  a  weak  attraction  between  the  chromosomes.  This  allows 
irregularities  of  distribution  in  the  heterotypic  mitosis,  so  that  both 
(unpaired)  chromosomes  belonging  to  one  pair  will  occasionally  enter 
the  same  daughter  nucleus  (see  p.  26).  Germ  cells  will  thus  arise, 
from  which  both  members  of  a  given  pair  of  chromosomes  are 
absent. 

4.  If  we  assume  qualitative  differences  between  the  chromosomes 
or  parts  of  them,  various  types  would  be  expected  to  originate  in  this 
manner,  each  of  them  lacking  the  ability  to  develop  certain  qualities 
possessed  by  the  parent  form.  On  this  view  the  mutations  of  Oeno¬ 
thera  Lamarckiana  are  an  instance  of  a  process  of  analysis  by  which 
from  the  parent  form  arises  a  series  of  types,  each  lacking  in  certain 
characters  or  capacities  possessed  by  the  parent.  This  hypothesis 
would  account  for  the  absence  of  reversions  arnong  Oenothera 
mutants,  and  perhaps  also  for  some  of  the  peculiarities  of  hybridiza¬ 
tion  in  Oenothera.  This  matter  will  be  considered  at  another  time. 
This  explanation  does  not  apply  to  all  the  mutants,  however;  for 
example,  O.  gigas. 


GATES— REDUCTION  IN  OENOTHERA 


31 


1908] 

5.  It  is  suggested  that  there  is  probably  a  direct  relation  between 
the  events  of  reduction  in  a  given  genus  and  its  variation,  as  well  as  its 
hybridization  phenomena. 

I  desire  to  express  my  thanks  to  Professors  John  M.  Coulter 
and  Charles  R.  Barnes  for  valuable  suggestions  and  adequate 
facilities  in  connection  with  this  work. 

The  University  oe  Chicago 

LITERATURE  CITED 

1.  Allen,  C.  E.,  Nuclear  division  in  the  pollen  mother  cells  of  Lilium  canadense. 
Annals  of  Botany  19:189-258.  pis.  6-g.  1905. 

2.  - ,  Das  Verhalten  der  Kernsubstanzen  wahrend  der  Synapsis  in  den 

Pollenmutterzellen  von  Lilium  canadense.  Jahrb.  Wiss.  Bot.  42:72-82. 
pi.  2.  1905. 

3.  Berghs,  Jules,  La  formation  des  chromosomes  heterotypiques  dans  murs  la 
sporogenese  vegetale.  I.  Depuis  le  spireme  jusqu’au  chromosomes  murs, 
dans  la  microsporogenese  d* Allium  fistulosum  et  Lilium  lancifolium  {speci- 
osum).  La  Cellule  21 : 173-189.  pi.  i.  1904. 

4.  - ,  II.  Depuis  la  sporogonie  jusqu’au  spireme  definitif,  dans  la  micro¬ 

sporogenese  de  I’d 21 : 383-397.  pi.  I.  1904. 

5.  - ,  III.  La  microsporogenese  de  Convallaria  maialis.  Idem  22:43-50. 

pi.  I.  1905. 

6.  - ,  IV.  La  microsporogenese  de  Drosera  rotundifolia,  Narthecium 

ossifragum,  et  Hellehorus  foetidus.  Idem  22:141-160.  pis.  2.  1905. 

7.  Cardiff,  Ira  D.,  A  study  of  synapsis  and  reduction.  Bull.  Torr.  Bot. 
Club  33:271-306.  pis.  12-15.  1906. 

8.  Farmer,  J.  B.,  and  Moore,  J.  E.  S.,  New  investigations  into  the  reduction 
phenomena  of  animals  and  plants.  Proc.  Roy.  Soc.  72 : 104-108.  figs.  6.  1903. 

9.  - - - — ,  On  the  maiotic  phase  (reduction  divisions)  in  animals  and  plants. 

Quart.  Jour.  Micr.  Sci.  48:489-557.  pis.  34-41.  1905. 

10.  Farmer,  J.  B.,  and  Shove,  Dorothy,  On  the  structure  and  development  of 
the  somatic  and  heterotype  chromosomes  of  Tradescantia  virginica.  Idem 

48:559-569-  pis.  42,  43’  1905- 

11.  Gates,  R.  R.,  Pollen  development  in  hybrids  of  Oenothera  lata  X  O. 
Lamarckiana,  and  its  relation  to  mutation.  Bot.  Gazette  43:81-115. 
pis.  2-4.  1907. 

12.  — - Hybridization  and  germ  cells  of  Oenothera  mutants.  Idem  44:1-21. 

figs.  3.  1907. 

13.  - ,  International  Zool.  Congress,  Boston,  Aug.,  1907. 

14.  - ,  The  chromosomes  of  Oenothera.  Science  N.  S.  27:193-195.  1908. 

15.  - — ,  Further  studies  on  the  chromosomes  of  Oenothera.  Idem  27:335. 


1908. 


32 


BOTANICAL  GAZETTE 


[JULY 


1 6.  Gregoire,  V.,  La  reduction  numerique  des  chromosomes  et  les  cineses  de 
maturation.  La  Cellule  21 : 297-314.  1904. 

17.  Lutz,  Anne  M.,  Chromosomes  of  the  somatic  cells  of  the  Oenotheras. 
Science  N.  S.  27:335.  1908. 

18.  Miyake,  Kiichi.  Ueber  Reduktionsteilung  in  den  Pollenmutterzellen 
einiger  Monokotylen.  Jahrb.  Wiss.  Bot.  42:83-120.  pis.  j-5.  1905. 

19.  Mother,  D.  M.,  The  development  of  the  heterotypic  chromosomes  in  pollen 
mother  cells.  Bot.  Gazette  40: 1 71-177.  1905. 

20.  - ,  The  development  of  the  heterotypic  chromosomes  in  pollen  mother 

cells.  Annals  of  Botany  21:309-347.  pis.  27,  28.  1907. 

21.  Nichols,  M.  Louise,  The  development  of  the  pollen  of  Sarracenia.  Bot. 
Gazette  45:31-37.  pi.  5.  1908. 

22.  Overton,  J.  B.,  Ueber  Reduktionsteilung  in  den  Pollenmutterzellen  einiger 
Dikotylen.  Jahrb.  Wiss.  Bot.  42:121-153.  pis.  6,  7.  1905. 

23.  Rosenberg,  O.,  Das  Verhalten  der  chromosomen  in  einer  hybriden  Pflanze. 
Ber.  Deutsch.  Bot.  Gesells.  21:110-119.  1903. 

24.  - ,  Ueber  die  Tetradentheilung  eines  Drosera-Bastardes.  Ber.  Deutsch. 

Bot.  Gesells.  22:47-53.  P^-  4-  1904- 

25.  - ,  Zur  Kenntniss  der  Reduktionstheilung  in  Pflanzen.  Bot.  Notiser 

1905:  pp.  24.  Jigs.  14. 

26.  - ,  Cytological  investigations  in  plant  hybrids.  Report  3d  Internat. 

Conf.  Genetics,  pp.  289-291.  1906. 

27.  - ,  Cytological  studies  on  the  apogamy  in  Hieracium.  Bot.  Tidsskrift 

28:143-170.  Jigs.  13.  pis.  7,  2.  1907. 

28.  ScHAEFNER,  J.  H.,  Chromosome  reduction  in  the-  microsporocytes  of 
Lilium  tigrinum.  Bot.  Gazette  41:183-191.  pis.  12,  13.  1906. 

29.  - ,  Synapsis  and  synizesis.  Ohio  Nat.  7:41-48.  pi.  4.  1907. 

30.  Strasburger,  E.,  Ueber  Reduktionstheilung.  Sitzungsber.  k.  k.  Preuss. 
Akad.  Wiss.  18:587-614.  Jigs.  13.  1904. 

31.  - ,  Typische  und  allotypische  Kerntheilung.  Jahrb.  Wiss.  Bot.  42:1- 

71.  pi.  I.  1905. 

32.  Tischler,  G.,  Zellstudien  an  sterilen  Bastardpflanzen.  Archiv  Zellforschung 
1:33-151-  fig^-  120.  1908. 

33.  Yamanouchi,  Shigeo,  Sporogenesis  in  Nephrodium.  Bot.  Gazette  45: 
1-30.  pis.  1-4.  1908. 

EXPLANATION  OF  PLATES  I-III 

The  figures  were  drawn  with  the  aid  of  a  Bausch  &  Lomb  camera  lucida.  All 
except  Jigs,  i  and  ig  were  drawn  under  a  Zeiss  apochromatic  objective  2™™  ap. 
1 . 30,  with  a  Zeiss  compensating  ocular  18.  The  figures  are  reduced  one-fourth  in 
reproduction,  giving  a  magnification  of  nearly  3000  diameters.  Fig.  i  was  drawn 
under  a  2™^  objective  and  compensating  ocular  6;  Jig.  ig  under  B.  &  L.  objective 
iV  N.  A.  1.32  and  Zeiss  ocular  18. 


GATES— REDUCTION  IN  OENOTHERA 


33 


1908] 

PLATE  1 

Figs,  i,  2. — Young  meristematic  cells  of  anther  primordium  showing  one 
large  nucleolus  and  several  smaller  ones,  and  chromatic  masses  adherent  to  the 
nuclear  membrane. 

Fig.  3. — Longitudinal  section  of  anther,  showing  size  relations  of  nucleoli  in 
sporogenous,  tapetal,  and  wall  cells. 

Fig.  4. — One  sporogenous  cell  from  stage  of  fig.  j,  previous  to  synapsis; 
cytoplasm  somewhat  vacuolate. 

Figs.  5,  7-9. — Nuclei  at  same  stage,  showing  fusions  of  nucleoli. 

Fig.  6. — Two  nucleoli  of  equal  size;  an  unusual  condition. 

Fig.  10. — Several  small  nucleoli,  and  no  indication  of  fusion. 

Fig.  II. — Nucleoli  of  young  pollen  grain  nucleus. 

Fig.  12. — Beginning  of  synaptic  contraction;  the  reticulum  has  contracted 
from  the  nuclear  membrane  on  all  sides,  leaving  several  loops  attached  to  the 
membrane;  on  the  side  on  which  the  reticulum  retains  the  curved  outline  of  the 
nuclear  membrane  the  latter  has  been  drawn  inward  attached  to  the  threads;  on 
the  rest  of  the  circumference,  between  the  loops,  the  nuclear  membrane  remains 
in  situ;  the  cytoplasm  is  perfectly  fixed. 

Fig.  13. — Another  contraction  stage,  showing  loops  attached  to  the  nuclear 
membrane,  which  is  intact. 

Fig.  14. — A  slightly  later  stage  of  contraction,  in  which  the  rearrangement 
of  threads  is  taking  place. 

Fig.  15. — Synapsis;  dark-staining  bodies  are  still  held  in  the  meshes  of  the 
spirem;  a  small  nu-cleolus,  usually  about  the  size  of  a  chromosome,  is  generally 
present  in  addition  to  the  large  nucleolus. 

Fig.  16. — After  synapsis;  the  thread  thicker  and  shorter  and  loosely  coiled. 

Fig.  17. — Slightly  later  stage  than  fig.  16,  and  less  deeply  stained;  thread 
shows  the  characteristic  light  and  dark  areas;  indications  of  parallel  threads  in 
two  places;  edge  of  thread  may  be  even  or  moniliform. — 5  ai. 

Fig.  18. — Later  stage;  thread  much  shortened  and  greatly  thickened  and 
entering  upon  second  contraction  phase;  nucleus  uncut. — 10  p. 

Fig.  19. — Higher  magnification  of  a  portion  of  the  thread  in  figs.  20  and  21. 

PLA  TE  II 

Fig.  20. — Second  contraction  stage;  a  pair  of  chromosomes  cut  off  from 
spirem;  nucleus  uncut. — 10  /x. 

Fig.  21. — Second  contraction  stage;  nucleus  uncut. 

Fig.  22. — Uncoiling  from  second  contraction  stage;  pair  of  chromosomes 
detached;  nucleus  uncut. 

Fig.  23. — Spirem  segmented  in  three  places,  each  segment  showing  constric¬ 
tions  which  will  form  the  chromosomes;  certain  chromosomes  already  detached; 
nucleus  uncut. 

Fig.  24. — Constriction  of  spirem  has  proceeded  farther,  the  chromosomes  being 
elongated  bodies  with  irregular  margins  like  the  spirem,  and  connected  by  rather 


34 


BOTANICAL  GAZETTE 


[JULY 


thick  “linin’’  bands;  pair  of  chromosomes  detached  earlier  lies  at  side  of  nucleus; 
n,  small  nucleolus;  nucleus  cut. 

Fig.  25. — Spirem  more  or  less  completely  segmented  into  chromosomes  while 
still  in  the  second  contraction  stage;  preparation  considerably  destained;  13 
chromosomes  in  view. 

Fig.  26. — Spirem  segmented,  showing  chain  of  eight  chromosomes  and  three 
pairs;  nucleus  uncut. 

Fig.  27. — Chain  of  six  chromosomes,  and  probably  four  pairs;  linin  con¬ 
nections  between  members  of  a  pair  not  always  visible;  nucleus  uncut. 

Fig.  28. — Fourteen  chromosomes;  several  small  nucleoli;  nucleus  uncut. 

Fig.  29. — Fourteen  chromosomes,  including  five  pairs  more  or  less  closely 
associated;  linin  connections  not  visible;  one  pair  of  chromosomes  has  already 
contracted  into  the  globular  shape. 

Fig.  30. — Fourteen  chromosomes,  several  in  pairs;  apparent  inequalities  in 
size  due  to  positions  in  which  some  of  the  chromosomes  are  lying. 

Fig.  31. — Slightly  later  stage;  the  fourteen  chromosomes  contracted  into  the 
globular  or  pear-shaped  definitive  form;  linin  connections  longer  and  extremely 
delicate;  nucleus  uncut. 

Figs.  32-34. — Other  groups  in  diakinesis,  showing  various  peculiarities  of 
chromosomes. 

Fig.  35. — Peculiar  case  of  spindle  formation;  three  nucleoli  present  and  four¬ 
teen  chromosomes,  including  three  or  four  pairs. 

Fig.  36. — Multipolar  stage  of  heterotypic  spindle;  two  more  or  less  closely 
united  pairs  of  chromosomes  present. 

PLA  TE  III 

Fig.  37. — Same  as  fig.  j6;  an  unusual  case  in  which  all  the  chromosomes 
are  closely  joined  in  pairs;  seven  such  pairs  present  and  a  small  nucleolus. 

Fig.  38. — Heterotypic  spindle  in  metaphase;  spindle  has  usually  more  mantle 
fibers  than  in  O.  Lamarckiana;  chromosomes  usually  loosely  arranged  in  equatorial 
region  of  spindle. 

Fig.  39. — Late  anaphase;  an  uncommon  case;  daughter  chromosomes  have 
failed  to  divide,  and  fibrillae  are  scattered  in  cytoplasm  at  side  of  cell;  chromatic 
staining  material  also  present. 

Fig.  40. — Telophase  of  heterotypic  mitosis;  exceptional  case,  in  which  a 
rather  sharp  pointed  spindle  is  formed  at  side  of  cell;  it  probably  originated  from 
the  fibrillae  shown  in  Jig.  jg. 

Fig.  41. — Early  anaphase  of  homotypic  mitosis;  small  nucleolus  having  the 
characteristic  appearance,  present  on  one  of  the  spindles. 

Fig.  42. — The  single  case  of  extra  nuclei  observed  in  O.  ruhrinervis  pollen 
mother  cells. 

Figs.  43,  44. — Nuclei  from  telophase  of  second  mitosis,  passing  into  resting 
condition. 

Figs.  45,  46. — Protoplasmic  connections  between  mother  cells. 


igo8] 


CzATES— -REDUCTION  IN  OENOTHERA 


33 


PLA  TE  1 

Figs,  i,  2. — Young  meristematic  cells  of  anther  primordium  showing  one 
large  nucleolus  and  several  smaller  ones,  and  chromatic  masses  adherent  to  the 
nuclear  membrane. 

Fig.  3.— Longitudinal  section  of  anther,  showing  size  relations  of  nucleoli  in 
sporogenous,  tapetal,  and  wall  cells. 

Fig.  4. — One  sporogenous  cell  from  stage  of  fig.  j,  previous  to  synapsis; 
cytoplasm  somewhat  vacuolate. 

Figs,  5,  7-9. — Nuclei  at  same  stage,  showing  fusions  of  nucleoli. 

Fig.  6. — Two  nucleoli  of  equal  size;  an  unusual  condition. 

Fig.  10, — Several  small  nucleoli,  and  no  indication  of  fusion. 

Fig.  II. — Nucleoli  of  young  pollen  grain  nucleus. 

Fig.  12. — Beginning  of  synaptic  contraction;  the  reticulum  has  contracted 
from  the  nuclear  membrane  on  all  sides,  leaving  several  loops  attached  to  the 
membrane;  on  the  side  on  which  the  reticulum  retains  the  curved  outline  of  the 
nuclear  membrane  the  latter  has  been  drawn  inward  attached  to  the  threads;  on 
the  rest  of  the  circumference,  between  the  loops,  the  nuclear  membrane  remains 
in  situ;  the  cytoplasm  is  perfectly  fixed. 

Fig.  13. — Another  contraction  stage,  showing  loops  attached  to  the  nuclear 
membrane,  which  is  intact. 

Fig.  14. — A  slightly  later  stage  of  contraction,  in  which  the  rearrangement 
of  threads  is  taking  place. 

Fig.  15. — Synapsis;  dark-staining  bodies  are  still  held  in  the  meshes  of  the 
spirem;  a  small  nucleolus,  usually  about  the  size  of  a  chromosome,  is  generally 
present  in  addition  to  the  large  nucleolus. 

Fig.  16. — After  synapsis;  the  thread  thicker  and  shorter  and  loosely  coiled. 

Fig.  17. — Slightly  later  stage  than  fig.  16,  and  less  deeply  stained;  thread 
shows  the  characteristic  light  and  dark  areas;  indications  of  parallel  threads  in 
two  places;  edge  of  thread  may  be  even  or  moniliform. — 5  u. 

Fig.  18. — Later  stage;  thread  much  shortened  and  greatly  thickened  and 
entering  upon  second  contraction  phase;  nucleus  uncut. — 10  u. 

Fig.  19. — Higher  magnification  of  a  portion  of  the  thread  in  figs.  20  and  21. 

PLATE  II 

Fig.  20. — Second  contraction  stage;  a  pair  of  chromosomes  cut  off  from 
spirem;  nucleus  uncut. — 10  u. 

Fig.  21. — Second  contraction  stage;  nucleus  uncut. 

Fig.  22. — Uncoiling  from  second  contraction  stage;  pair  of  chromosomes 
detached;  nucleus  uncut. 

Fig.  23. — Spirem  segmented  in  three  places,  each  segment  showing  constric¬ 
tions  which  will  form  the  chromosomes;  certain  chromosomes  already  detached; 
nucleus  uncut. 

Fig.  24. — Constriction  of  spirem  has  proceeded  farther,  the  chromosomes  being 
elongated  bodies  with  irregular  margins  like  the  spirem,  and  connected  by  rather 


34 


BOTANICAL  GAZETTE 


[JULY 


thick  “linin’’  bands;  pair  of  chromosomes  detached  earlier  lies  at  side  of  nucleus; 
n,  small  nucleolus;  nucleus  cut. 

Fig.  25.^ — Spirem  more  or  less  completely  segmented  into  chromosomes  while 
still  in  the  second  contraction  stage;  preparation  considerably  destained;  13 
chromosomes  in  view. 

Fig.  26. — Spirem  segmented,  showing  chain  of  eight  chromosomes  and  three 
pairs;  nucleus  uncut. 

Fig.  27. — Chain  of  six  chromosomes,  and  probably  four  pairs;  linin  con¬ 
nections  between  members  of  a  pair  not  always  visible;  nucleus  uncut. 

Fig.  28. — Fourteen  chromosomes;  several  small  nucleoli;  nucleus  uncut. 

Fig.  29. — Fourteen  chromosomes,  including  five  pairs  more  or  less  closely 
associated;  linin  connections  not  visible;  one  pair  of  chromosomes  has  already 
contracted  into  the  globular  shape. 

Fig.  30. — Fourteen  chromosomes,  several  in  pairs;  apparent  inequalities  in 
size  due  to  positions  in  which  some  of  the  chromosomes  are  lying. 

Fig.  31. — Slightly  later  stage;  the  fourteen  chromosomes  contracted  into  the 
globular  or  pear-shaped  definitive  form;  linin  connections  longer  and  extremely 
delicate;  nucleus  uncut. 

Figs.  32-34. — Other  groups  in  diakinesis,  showing  various  peculiarities  of 
chromosomes. 

Fig.  35. — Peculiar  case  of  spindle  formation;  three  nucleoli  present  and  four¬ 
teen  chromosomes,  including  three  or  four  pairs. 

Fig.  36.— Multipolar  stage  of  heterotypic  spindle;  two  more  or  less  closely 
united  pairs  of  chromosomes  present. 

PLA  TE  III 

Fig.  37. — Same  as  jig.  j6;  an  unusual  case  in  which  all  the  chromosomes 
are  closely  joined  in  pairs;  seven  such  pairs  present  and  a  small  nucleolus. 

Fig.  38. — Heterotypic  spindle  in  metaphase;  spindle  has  usually  more  mantle 
fibers  than  in  O.  Lamarckiana;  chromosomes  usually  loosely  arranged  in  equatorial 
region  of  spindle. 

Fig.  39. — Late  anaphase;  an  uncommon  case;  daughter  chromosomes  have 
failed  to  divide,  and  fibrillae  are  scattered  in  cytoplasm  at  side  of  cell;  chromatic 
staining  material  also  present. 

Fig.  40. — Telophase  of  heterotypic  mitosis;  exceptional  case,  in  which  a 
rather  sharp  pointed  spindle  is  formed  at  side  of  cell;  it  probably  originated  from 
the  fibrillae  shown  in  fig.  jp. 

Fig.  41.— Early  anaphase  of  homotypic  mitosis;  small  nucleolus  having  the 
characteristic  appearance,  present  on  one  of  the  spindles. 

Fig.  42. — The  single  case  of  extra  nuclei  observed  in  O.  ruhrinervis  pollen 
mother  cells. 

Figs.  43,  44. — Nuclei  from  telophase  of  second  mitosis,  passing  into  resting 
condition. 

Figs.  45,  46. — Protoplasmic  connections  between  mother  cells. 


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